Luiziana Ferreira da SilvaLab of BioproductsDepartment of MicrobiologyInstitute of Biomedical SciencesUniversity of Satildeo Paulo - Brazil

Microbial GeneticsBiotechnology ndash Microbial anticancer compounds

Nitrogen Biological Fixation in sugarcane

Bacterial MetabolismBacterial products ndash biopolymers biosurfactants

Bacterial MetabolismBiotechnology ndashbacterial biopolymers

Microbial ecology Plant-microorganism interactionsBiotechnology

Harmfull effects Human

Animals

Agriculture

Environment

BenefitsMedicine

Pharmaceutical industry

Food Industry

Agriculture

Environment

Roles of microbiomes

Studies in humansSkin protection

Obesity-associated gut microbiome

Brain development amp behavior

Faecal bacteriotherapy in the treatment of recurrent C dificilediarrhea

Roles to plant and animals

A number of new surveys are helping scientists understand the many ecosystems our bodies offer to microbes and their interactions

A relevant part of our society is still unaware about the benefits and the contribution to our lives that were achieved because of the action of microorganisms

Country extension

Various climates

Different environments

Different crops cultivated

Variable and heterogeneous natural microbiota

New species discovered

New genes

Generation of bioproducts

Platforms for alternative products or alternative routes

Nitrogen fixing bacteria

Antitumoral agents

Endophytes

Biopolymers

1986 isolated from Brazilian Savannah (cerrado) a

soil showing high acidity low levels of N and organic matter ndashBarbosa et al 1986 Barbosa amp Carvalhal 1988

Physiologic characteristics related to the environment were studied in this bacterium

bull Nitrogen fixing ability under adverse conditions low pH and underhigh concentrations of toxic compounds

bull Role of exopolysaccharide in protecting the nitrogenase from oxygendeleterious effects

bull Stimulation of other bacteria in N-free mediumbull Liberation of aminoacids in N-free culture mediumbull Cyanophicin like intracelular reserves

DrHeloiza R Barbosa

B derxii pure amp associate cultures

E coli pure amp associate cultures

Stimulating both heteroand autothrophs usuallypresent in soil

Nitrogenated reserve granulesCyanophycin-likeUnder study

Biotechnological applicationsPurified granules can be chemically converted to a derivative with reduced arginine content or to completely biodegradable poly(aspartic acid) which can be used as a substitute for nonbiodegradable polyacrylateswith many technical and medical applications

Production of antitumoral agents

Polyketydes (PKS)

DrGabriel Padilla

Gonccedilalves de Lima et al 1969Retamycin

Anthracyclines are active against bull breast cancerbull lymphomas bull acute leukemiasbull neuroblastomasbull bone and soft-tissue

sarcomas

Studies on the mechanism of action

Chemical structures of cosmomycinsproduced by Streptomyces olindensis

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

Microbial GeneticsBiotechnology ndash Microbial anticancer compounds

Nitrogen Biological Fixation in sugarcane

Bacterial MetabolismBacterial products ndash biopolymers biosurfactants

Bacterial MetabolismBiotechnology ndashbacterial biopolymers

Microbial ecology Plant-microorganism interactionsBiotechnology

Harmfull effects Human

Animals

Agriculture

Environment

BenefitsMedicine

Pharmaceutical industry

Food Industry

Agriculture

Environment

Roles of microbiomes

Studies in humansSkin protection

Obesity-associated gut microbiome

Brain development amp behavior

Faecal bacteriotherapy in the treatment of recurrent C dificilediarrhea

Roles to plant and animals

A number of new surveys are helping scientists understand the many ecosystems our bodies offer to microbes and their interactions

A relevant part of our society is still unaware about the benefits and the contribution to our lives that were achieved because of the action of microorganisms

Country extension

Various climates

Different environments

Different crops cultivated

Variable and heterogeneous natural microbiota

New species discovered

New genes

Generation of bioproducts

Platforms for alternative products or alternative routes

Nitrogen fixing bacteria

Antitumoral agents

Endophytes

Biopolymers

1986 isolated from Brazilian Savannah (cerrado) a

soil showing high acidity low levels of N and organic matter ndashBarbosa et al 1986 Barbosa amp Carvalhal 1988

Physiologic characteristics related to the environment were studied in this bacterium

bull Nitrogen fixing ability under adverse conditions low pH and underhigh concentrations of toxic compounds

bull Role of exopolysaccharide in protecting the nitrogenase from oxygendeleterious effects

bull Stimulation of other bacteria in N-free mediumbull Liberation of aminoacids in N-free culture mediumbull Cyanophicin like intracelular reserves

DrHeloiza R Barbosa

B derxii pure amp associate cultures

E coli pure amp associate cultures

Stimulating both heteroand autothrophs usuallypresent in soil

Nitrogenated reserve granulesCyanophycin-likeUnder study

Biotechnological applicationsPurified granules can be chemically converted to a derivative with reduced arginine content or to completely biodegradable poly(aspartic acid) which can be used as a substitute for nonbiodegradable polyacrylateswith many technical and medical applications

Production of antitumoral agents

Polyketydes (PKS)

DrGabriel Padilla

Gonccedilalves de Lima et al 1969Retamycin

Anthracyclines are active against bull breast cancerbull lymphomas bull acute leukemiasbull neuroblastomasbull bone and soft-tissue

sarcomas

Studies on the mechanism of action

Chemical structures of cosmomycinsproduced by Streptomyces olindensis

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

Harmfull effects Human

Animals

Agriculture

Environment

BenefitsMedicine

Pharmaceutical industry

Food Industry

Agriculture

Environment

Roles of microbiomes

Studies in humansSkin protection

Obesity-associated gut microbiome

Brain development amp behavior

Faecal bacteriotherapy in the treatment of recurrent C dificilediarrhea

Roles to plant and animals

A number of new surveys are helping scientists understand the many ecosystems our bodies offer to microbes and their interactions

A relevant part of our society is still unaware about the benefits and the contribution to our lives that were achieved because of the action of microorganisms

Country extension

Various climates

Different environments

Different crops cultivated

Variable and heterogeneous natural microbiota

New species discovered

New genes

Generation of bioproducts

Platforms for alternative products or alternative routes

Nitrogen fixing bacteria

Antitumoral agents

Endophytes

Biopolymers

1986 isolated from Brazilian Savannah (cerrado) a

soil showing high acidity low levels of N and organic matter ndashBarbosa et al 1986 Barbosa amp Carvalhal 1988

Physiologic characteristics related to the environment were studied in this bacterium

bull Nitrogen fixing ability under adverse conditions low pH and underhigh concentrations of toxic compounds

bull Role of exopolysaccharide in protecting the nitrogenase from oxygendeleterious effects

bull Stimulation of other bacteria in N-free mediumbull Liberation of aminoacids in N-free culture mediumbull Cyanophicin like intracelular reserves

DrHeloiza R Barbosa

B derxii pure amp associate cultures

E coli pure amp associate cultures

Stimulating both heteroand autothrophs usuallypresent in soil

Nitrogenated reserve granulesCyanophycin-likeUnder study

Biotechnological applicationsPurified granules can be chemically converted to a derivative with reduced arginine content or to completely biodegradable poly(aspartic acid) which can be used as a substitute for nonbiodegradable polyacrylateswith many technical and medical applications

Production of antitumoral agents

Polyketydes (PKS)

DrGabriel Padilla

Gonccedilalves de Lima et al 1969Retamycin

Anthracyclines are active against bull breast cancerbull lymphomas bull acute leukemiasbull neuroblastomasbull bone and soft-tissue

sarcomas

Studies on the mechanism of action

Chemical structures of cosmomycinsproduced by Streptomyces olindensis

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

Roles of microbiomes

Studies in humansSkin protection

Obesity-associated gut microbiome

Brain development amp behavior

Faecal bacteriotherapy in the treatment of recurrent C dificilediarrhea

Roles to plant and animals

A number of new surveys are helping scientists understand the many ecosystems our bodies offer to microbes and their interactions

A relevant part of our society is still unaware about the benefits and the contribution to our lives that were achieved because of the action of microorganisms

Country extension

Various climates

Different environments

Different crops cultivated

Variable and heterogeneous natural microbiota

New species discovered

New genes

Generation of bioproducts

Platforms for alternative products or alternative routes

Nitrogen fixing bacteria

Antitumoral agents

Endophytes

Biopolymers

1986 isolated from Brazilian Savannah (cerrado) a

soil showing high acidity low levels of N and organic matter ndashBarbosa et al 1986 Barbosa amp Carvalhal 1988

Physiologic characteristics related to the environment were studied in this bacterium

bull Nitrogen fixing ability under adverse conditions low pH and underhigh concentrations of toxic compounds

bull Role of exopolysaccharide in protecting the nitrogenase from oxygendeleterious effects

bull Stimulation of other bacteria in N-free mediumbull Liberation of aminoacids in N-free culture mediumbull Cyanophicin like intracelular reserves

DrHeloiza R Barbosa

B derxii pure amp associate cultures

E coli pure amp associate cultures

Stimulating both heteroand autothrophs usuallypresent in soil

Nitrogenated reserve granulesCyanophycin-likeUnder study

Biotechnological applicationsPurified granules can be chemically converted to a derivative with reduced arginine content or to completely biodegradable poly(aspartic acid) which can be used as a substitute for nonbiodegradable polyacrylateswith many technical and medical applications

Production of antitumoral agents

Polyketydes (PKS)

DrGabriel Padilla

Gonccedilalves de Lima et al 1969Retamycin

Anthracyclines are active against bull breast cancerbull lymphomas bull acute leukemiasbull neuroblastomasbull bone and soft-tissue

sarcomas

Studies on the mechanism of action

Chemical structures of cosmomycinsproduced by Streptomyces olindensis

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

A relevant part of our society is still unaware about the benefits and the contribution to our lives that were achieved because of the action of microorganisms

Country extension

Various climates

Different environments

Different crops cultivated

Variable and heterogeneous natural microbiota

New species discovered

New genes

Generation of bioproducts

Platforms for alternative products or alternative routes

Nitrogen fixing bacteria

Antitumoral agents

Endophytes

Biopolymers

1986 isolated from Brazilian Savannah (cerrado) a

soil showing high acidity low levels of N and organic matter ndashBarbosa et al 1986 Barbosa amp Carvalhal 1988

Physiologic characteristics related to the environment were studied in this bacterium

bull Nitrogen fixing ability under adverse conditions low pH and underhigh concentrations of toxic compounds

bull Role of exopolysaccharide in protecting the nitrogenase from oxygendeleterious effects

bull Stimulation of other bacteria in N-free mediumbull Liberation of aminoacids in N-free culture mediumbull Cyanophicin like intracelular reserves

DrHeloiza R Barbosa

B derxii pure amp associate cultures

E coli pure amp associate cultures

Stimulating both heteroand autothrophs usuallypresent in soil

Nitrogenated reserve granulesCyanophycin-likeUnder study

Biotechnological applicationsPurified granules can be chemically converted to a derivative with reduced arginine content or to completely biodegradable poly(aspartic acid) which can be used as a substitute for nonbiodegradable polyacrylateswith many technical and medical applications

Production of antitumoral agents

Polyketydes (PKS)

DrGabriel Padilla

Gonccedilalves de Lima et al 1969Retamycin

Anthracyclines are active against bull breast cancerbull lymphomas bull acute leukemiasbull neuroblastomasbull bone and soft-tissue

sarcomas

Studies on the mechanism of action

Chemical structures of cosmomycinsproduced by Streptomyces olindensis

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

Country extension

Various climates

Different environments

Different crops cultivated

Variable and heterogeneous natural microbiota

New species discovered

New genes

Generation of bioproducts

Platforms for alternative products or alternative routes

Nitrogen fixing bacteria

Antitumoral agents

Endophytes

Biopolymers

1986 isolated from Brazilian Savannah (cerrado) a

soil showing high acidity low levels of N and organic matter ndashBarbosa et al 1986 Barbosa amp Carvalhal 1988

Physiologic characteristics related to the environment were studied in this bacterium

bull Nitrogen fixing ability under adverse conditions low pH and underhigh concentrations of toxic compounds

bull Role of exopolysaccharide in protecting the nitrogenase from oxygendeleterious effects

bull Stimulation of other bacteria in N-free mediumbull Liberation of aminoacids in N-free culture mediumbull Cyanophicin like intracelular reserves

DrHeloiza R Barbosa

B derxii pure amp associate cultures

E coli pure amp associate cultures

Stimulating both heteroand autothrophs usuallypresent in soil

Nitrogenated reserve granulesCyanophycin-likeUnder study

Biotechnological applicationsPurified granules can be chemically converted to a derivative with reduced arginine content or to completely biodegradable poly(aspartic acid) which can be used as a substitute for nonbiodegradable polyacrylateswith many technical and medical applications

Production of antitumoral agents

Polyketydes (PKS)

DrGabriel Padilla

Gonccedilalves de Lima et al 1969Retamycin

Anthracyclines are active against bull breast cancerbull lymphomas bull acute leukemiasbull neuroblastomasbull bone and soft-tissue

sarcomas

Studies on the mechanism of action

Chemical structures of cosmomycinsproduced by Streptomyces olindensis

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

New species discovered

New genes

Generation of bioproducts

Platforms for alternative products or alternative routes

Nitrogen fixing bacteria

Antitumoral agents

Endophytes

Biopolymers

1986 isolated from Brazilian Savannah (cerrado) a

soil showing high acidity low levels of N and organic matter ndashBarbosa et al 1986 Barbosa amp Carvalhal 1988

Physiologic characteristics related to the environment were studied in this bacterium

bull Nitrogen fixing ability under adverse conditions low pH and underhigh concentrations of toxic compounds

bull Role of exopolysaccharide in protecting the nitrogenase from oxygendeleterious effects

bull Stimulation of other bacteria in N-free mediumbull Liberation of aminoacids in N-free culture mediumbull Cyanophicin like intracelular reserves

DrHeloiza R Barbosa

B derxii pure amp associate cultures

E coli pure amp associate cultures

Stimulating both heteroand autothrophs usuallypresent in soil

Nitrogenated reserve granulesCyanophycin-likeUnder study

Biotechnological applicationsPurified granules can be chemically converted to a derivative with reduced arginine content or to completely biodegradable poly(aspartic acid) which can be used as a substitute for nonbiodegradable polyacrylateswith many technical and medical applications

Production of antitumoral agents

Polyketydes (PKS)

DrGabriel Padilla

Gonccedilalves de Lima et al 1969Retamycin

Anthracyclines are active against bull breast cancerbull lymphomas bull acute leukemiasbull neuroblastomasbull bone and soft-tissue

sarcomas

Studies on the mechanism of action

Chemical structures of cosmomycinsproduced by Streptomyces olindensis

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

Nitrogen fixing bacteria

Antitumoral agents

Endophytes

Biopolymers

1986 isolated from Brazilian Savannah (cerrado) a

soil showing high acidity low levels of N and organic matter ndashBarbosa et al 1986 Barbosa amp Carvalhal 1988

Physiologic characteristics related to the environment were studied in this bacterium

bull Nitrogen fixing ability under adverse conditions low pH and underhigh concentrations of toxic compounds

bull Role of exopolysaccharide in protecting the nitrogenase from oxygendeleterious effects

bull Stimulation of other bacteria in N-free mediumbull Liberation of aminoacids in N-free culture mediumbull Cyanophicin like intracelular reserves

DrHeloiza R Barbosa

B derxii pure amp associate cultures

E coli pure amp associate cultures

Stimulating both heteroand autothrophs usuallypresent in soil

Nitrogenated reserve granulesCyanophycin-likeUnder study

Biotechnological applicationsPurified granules can be chemically converted to a derivative with reduced arginine content or to completely biodegradable poly(aspartic acid) which can be used as a substitute for nonbiodegradable polyacrylateswith many technical and medical applications

Production of antitumoral agents

Polyketydes (PKS)

DrGabriel Padilla

Gonccedilalves de Lima et al 1969Retamycin

Anthracyclines are active against bull breast cancerbull lymphomas bull acute leukemiasbull neuroblastomasbull bone and soft-tissue

sarcomas

Studies on the mechanism of action

Chemical structures of cosmomycinsproduced by Streptomyces olindensis

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

1986 isolated from Brazilian Savannah (cerrado) a

soil showing high acidity low levels of N and organic matter ndashBarbosa et al 1986 Barbosa amp Carvalhal 1988

Physiologic characteristics related to the environment were studied in this bacterium

bull Nitrogen fixing ability under adverse conditions low pH and underhigh concentrations of toxic compounds

bull Role of exopolysaccharide in protecting the nitrogenase from oxygendeleterious effects

bull Stimulation of other bacteria in N-free mediumbull Liberation of aminoacids in N-free culture mediumbull Cyanophicin like intracelular reserves

DrHeloiza R Barbosa

B derxii pure amp associate cultures

E coli pure amp associate cultures

Stimulating both heteroand autothrophs usuallypresent in soil

Nitrogenated reserve granulesCyanophycin-likeUnder study

Biotechnological applicationsPurified granules can be chemically converted to a derivative with reduced arginine content or to completely biodegradable poly(aspartic acid) which can be used as a substitute for nonbiodegradable polyacrylateswith many technical and medical applications

Production of antitumoral agents

Polyketydes (PKS)

DrGabriel Padilla

Gonccedilalves de Lima et al 1969Retamycin

Anthracyclines are active against bull breast cancerbull lymphomas bull acute leukemiasbull neuroblastomasbull bone and soft-tissue

sarcomas

Studies on the mechanism of action

Chemical structures of cosmomycinsproduced by Streptomyces olindensis

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

B derxii pure amp associate cultures

E coli pure amp associate cultures

Stimulating both heteroand autothrophs usuallypresent in soil

Nitrogenated reserve granulesCyanophycin-likeUnder study

Biotechnological applicationsPurified granules can be chemically converted to a derivative with reduced arginine content or to completely biodegradable poly(aspartic acid) which can be used as a substitute for nonbiodegradable polyacrylateswith many technical and medical applications

Production of antitumoral agents

Polyketydes (PKS)

DrGabriel Padilla

Gonccedilalves de Lima et al 1969Retamycin

Anthracyclines are active against bull breast cancerbull lymphomas bull acute leukemiasbull neuroblastomasbull bone and soft-tissue

sarcomas

Studies on the mechanism of action

Chemical structures of cosmomycinsproduced by Streptomyces olindensis

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

Nitrogenated reserve granulesCyanophycin-likeUnder study

Biotechnological applicationsPurified granules can be chemically converted to a derivative with reduced arginine content or to completely biodegradable poly(aspartic acid) which can be used as a substitute for nonbiodegradable polyacrylateswith many technical and medical applications

Production of antitumoral agents

Polyketydes (PKS)

DrGabriel Padilla

Gonccedilalves de Lima et al 1969Retamycin

Anthracyclines are active against bull breast cancerbull lymphomas bull acute leukemiasbull neuroblastomasbull bone and soft-tissue

sarcomas

Studies on the mechanism of action

Chemical structures of cosmomycinsproduced by Streptomyces olindensis

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

Production of antitumoral agents

Polyketydes (PKS)

DrGabriel Padilla

Gonccedilalves de Lima et al 1969Retamycin

Anthracyclines are active against bull breast cancerbull lymphomas bull acute leukemiasbull neuroblastomasbull bone and soft-tissue

sarcomas

Studies on the mechanism of action

Chemical structures of cosmomycinsproduced by Streptomyces olindensis

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

Studies on the mechanism of action

Chemical structures of cosmomycinsproduced by Streptomyces olindensis

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

Chemical structures of cosmomycinsproduced by Streptomyces olindensis

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

Garrido LM1 Lomboacute F Baig I Nur-E-Alam M Furlan RL Borda CC Brantildea A Meacutendez C Salas JA Rohr J Padilla G

Genetic organization of the sequenced DNA region and constructs used for the generation of the different mutants The black triangle indicates the apramycinresistance cassette Restriction sites indicated with an asterisk are not unique sites SaSau3AI NcNcoI BaBamHI NrNruI

Chemical structures of compounds isolated from Streptomyces olindensisΔcosK (a) and S olindensisΔcosG (b)

Proposed pathway for glycosylation events in cosmomycin D biosynthesis

DrWelington L Araujo

Characterization of mutants

a-1

a-2

a-3

b-1

b-2

c-1

c-2

d e

f

g

h

Random mutagenesis to identify genes related to biosynthetic pathway

KS AT P

P TEPKSi 10

2069 aa

KS AT DH ER KR P

P

PKSi 5

2585 aa

KS AT TEP

P

PKSi 11

2037 aa

KS AT P

P

P

P M H PPKSi 12

2637 aa

KS ATD

HM KR P

P C A RP

P

PKS-NRPS (PKSi6)

3981 aa

K

S

PKSIII

406 aa

PKSi 2KS AT M (ER)

K

RP

P

D

H

2262 aa

KS ATD

HM KR P

P

PKSi 1

2515 aa

KS AT DH MD

HKR P

P

PKSi 3

2576 aa

P

PKS ATD

HKR

PKSi 4

1777 aa

KS ATP

P

P

P TEPKSi 7

2145 aa

P

PKS AT DH M ER KRPKSi 8

2488 aa

P

PKS AT M RPKSi 9

2576 aa

K

S

A

T

K

R

D

H

E

R

T

E

M

P

P

H

P

R

Ketosynthase domain

Acyl transferase domain

Ketoreductase domain

Dehidrathese domain

Enoil reductase domain

Thioesterase domain

Methyltransferase domain

Acyl Carrier protrein domain

Hydrolase domain

Acyl CoA reductase domain

Ketosynthase domain

13 hypothetical PKS clusters identified in the E nigrum P16 genome Aminoacid numbers and domain positions are in scale

Epicolactone

New metabolite from Epicoccum nigrum

Associated to microbial control

Two PKS regulators were identified

Identification of the PKS gene cluster

Dr J Gregoacuterio C Gomez Dr Luiziana F Silva

PHA are accumulated as intracelular granules by bacteria

C4 amp C5

Short-chain length monomers

PHA scl

C6 amp C12

Medium-chain length

monomers PHA mcl

Naturally PHA-accumulating bacteria produce eitherPHAscl or PHAmcl in large amounts

Monomer composition is responsible for PHA properties and applications

730

Features influencing PHA monomer composition

Isolation amp evaluation of soil bacteria on the ability to produce PHB

from Sucrose Glucose and Fructose

Table 2 Production of P(3HB-co-3HV) from glucose plus propionic acid

Residual PHA

Strains CDWc

(gL)Carbohydrates

()

CDWc3HB

mol

3HV

molY3HVPrp

b

A eutrophus DSM 54 392 00 714 961 39 013

A latus DSM 1123 095 1018 146 550 450 007

Pcepacia DSM 50181 335 19 384 973 27 004

IPT-040 377 17 323 971 29 005

IPT-044 392 17 511 971 29 007

IPT-045 373 00 494 962 38 008

IPT-048 297 00 443 962 38 006

IPT-055 427 722 15 1000 00 000

IPT-056 360 313 309 985 15 002

IPT-076 506 19 568 971 29 010

IPT-083 489 52 568 969 31 010

IPT-086a 206 75 390 899 101 009

IPT-098 590 00 177 947 53 007

IPT-101 298 418 323 954 46 005

Gomez et al 1996

Improvement to

incorporate 3HV units

Strain Phenotype

strategy

3HB

mol

3HV mol

Y3HVprp

gg

B sacchari wild type 938 62 010

189 prp UV mutant 436 564 090

189 Sucrosepropionatefeeding rates

920 180 127

Y3HVprp = 3HV yield from propionic acid

Maximum theoretical yield = 135 gg

P3HB-co-3HV from sucrose and propionic acid

3HB C4-monomer PHA

3HV ndash C5

Inactivation on the propionate

catabolic pathway

2MCC = 2 methyl citrate cycle

prpR

ORF1 ORF3 ORF4 ORF5ORF6 ORF7 ORF8 ORF9ORF2

prpB prpC acnM

EcoRIEcoRI EcoRIEcoRI

EcoRI

SalISalI

4 kbp 29 kbp 2 kbp 9 kbp

A

B

putative lipoprotein

52 bp

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

Proposed pathway for glycosylation events in cosmomycin D biosynthesis

DrWelington L Araujo

Characterization of mutants

a-1

a-2

a-3

b-1

b-2

c-1

c-2

d e

f

g

h

Random mutagenesis to identify genes related to biosynthetic pathway

KS AT P

P TEPKSi 10

2069 aa

KS AT DH ER KR P

P

PKSi 5

2585 aa

KS AT TEP

P

PKSi 11

2037 aa

KS AT P

P

P

P M H PPKSi 12

2637 aa

KS ATD

HM KR P

P C A RP

P

PKS-NRPS (PKSi6)

3981 aa

K

S

PKSIII

406 aa

PKSi 2KS AT M (ER)

K

RP

P

D

H

2262 aa

KS ATD

HM KR P

P

PKSi 1

2515 aa

KS AT DH MD

HKR P

P

PKSi 3

2576 aa

P

PKS ATD

HKR

PKSi 4

1777 aa

KS ATP

P

P

P TEPKSi 7

2145 aa

P

PKS AT DH M ER KRPKSi 8

2488 aa

P

PKS AT M RPKSi 9

2576 aa

K

S

A

T

K

R

D

H

E

R

T

E

M

P

P

H

P

R

Ketosynthase domain

Acyl transferase domain

Ketoreductase domain

Dehidrathese domain

Enoil reductase domain

Thioesterase domain

Methyltransferase domain

Acyl Carrier protrein domain

Hydrolase domain

Acyl CoA reductase domain

Ketosynthase domain

13 hypothetical PKS clusters identified in the E nigrum P16 genome Aminoacid numbers and domain positions are in scale

Epicolactone

New metabolite from Epicoccum nigrum

Associated to microbial control

Two PKS regulators were identified

Identification of the PKS gene cluster

Dr J Gregoacuterio C Gomez Dr Luiziana F Silva

PHA are accumulated as intracelular granules by bacteria

C4 amp C5

Short-chain length monomers

PHA scl

C6 amp C12

Medium-chain length

monomers PHA mcl

Naturally PHA-accumulating bacteria produce eitherPHAscl or PHAmcl in large amounts

Monomer composition is responsible for PHA properties and applications

730

Features influencing PHA monomer composition

Isolation amp evaluation of soil bacteria on the ability to produce PHB

from Sucrose Glucose and Fructose

Table 2 Production of P(3HB-co-3HV) from glucose plus propionic acid

Residual PHA

Strains CDWc

(gL)Carbohydrates

()

CDWc3HB

mol

3HV

molY3HVPrp

b

A eutrophus DSM 54 392 00 714 961 39 013

A latus DSM 1123 095 1018 146 550 450 007

Pcepacia DSM 50181 335 19 384 973 27 004

IPT-040 377 17 323 971 29 005

IPT-044 392 17 511 971 29 007

IPT-045 373 00 494 962 38 008

IPT-048 297 00 443 962 38 006

IPT-055 427 722 15 1000 00 000

IPT-056 360 313 309 985 15 002

IPT-076 506 19 568 971 29 010

IPT-083 489 52 568 969 31 010

IPT-086a 206 75 390 899 101 009

IPT-098 590 00 177 947 53 007

IPT-101 298 418 323 954 46 005

Gomez et al 1996

Improvement to

incorporate 3HV units

Strain Phenotype

strategy

3HB

mol

3HV mol

Y3HVprp

gg

B sacchari wild type 938 62 010

189 prp UV mutant 436 564 090

189 Sucrosepropionatefeeding rates

920 180 127

Y3HVprp = 3HV yield from propionic acid

Maximum theoretical yield = 135 gg

P3HB-co-3HV from sucrose and propionic acid

3HB C4-monomer PHA

3HV ndash C5

Inactivation on the propionate

catabolic pathway

2MCC = 2 methyl citrate cycle

prpR

ORF1 ORF3 ORF4 ORF5ORF6 ORF7 ORF8 ORF9ORF2

prpB prpC acnM

EcoRIEcoRI EcoRIEcoRI

EcoRI

SalISalI

4 kbp 29 kbp 2 kbp 9 kbp

A

B

putative lipoprotein

52 bp

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

DrWelington L Araujo

Characterization of mutants

a-1

a-2

a-3

b-1

b-2

c-1

c-2

d e

f

g

h

Random mutagenesis to identify genes related to biosynthetic pathway

KS AT P

P TEPKSi 10

2069 aa

KS AT DH ER KR P

P

PKSi 5

2585 aa

KS AT TEP

P

PKSi 11

2037 aa

KS AT P

P

P

P M H PPKSi 12

2637 aa

KS ATD

HM KR P

P C A RP

P

PKS-NRPS (PKSi6)

3981 aa

K

S

PKSIII

406 aa

PKSi 2KS AT M (ER)

K

RP

P

D

H

2262 aa

KS ATD

HM KR P

P

PKSi 1

2515 aa

KS AT DH MD

HKR P

P

PKSi 3

2576 aa

P

PKS ATD

HKR

PKSi 4

1777 aa

KS ATP

P

P

P TEPKSi 7

2145 aa

P

PKS AT DH M ER KRPKSi 8

2488 aa

P

PKS AT M RPKSi 9

2576 aa

K

S

A

T

K

R

D

H

E

R

T

E

M

P

P

H

P

R

Ketosynthase domain

Acyl transferase domain

Ketoreductase domain

Dehidrathese domain

Enoil reductase domain

Thioesterase domain

Methyltransferase domain

Acyl Carrier protrein domain

Hydrolase domain

Acyl CoA reductase domain

Ketosynthase domain

13 hypothetical PKS clusters identified in the E nigrum P16 genome Aminoacid numbers and domain positions are in scale

Epicolactone

New metabolite from Epicoccum nigrum

Associated to microbial control

Two PKS regulators were identified

Identification of the PKS gene cluster

Dr J Gregoacuterio C Gomez Dr Luiziana F Silva

PHA are accumulated as intracelular granules by bacteria

C4 amp C5

Short-chain length monomers

PHA scl

C6 amp C12

Medium-chain length

monomers PHA mcl

Naturally PHA-accumulating bacteria produce eitherPHAscl or PHAmcl in large amounts

Monomer composition is responsible for PHA properties and applications

730

Features influencing PHA monomer composition

Isolation amp evaluation of soil bacteria on the ability to produce PHB

from Sucrose Glucose and Fructose

Table 2 Production of P(3HB-co-3HV) from glucose plus propionic acid

Residual PHA

Strains CDWc

(gL)Carbohydrates

()

CDWc3HB

mol

3HV

molY3HVPrp

b

A eutrophus DSM 54 392 00 714 961 39 013

A latus DSM 1123 095 1018 146 550 450 007

Pcepacia DSM 50181 335 19 384 973 27 004

IPT-040 377 17 323 971 29 005

IPT-044 392 17 511 971 29 007

IPT-045 373 00 494 962 38 008

IPT-048 297 00 443 962 38 006

IPT-055 427 722 15 1000 00 000

IPT-056 360 313 309 985 15 002

IPT-076 506 19 568 971 29 010

IPT-083 489 52 568 969 31 010

IPT-086a 206 75 390 899 101 009

IPT-098 590 00 177 947 53 007

IPT-101 298 418 323 954 46 005

Gomez et al 1996

Improvement to

incorporate 3HV units

Strain Phenotype

strategy

3HB

mol

3HV mol

Y3HVprp

gg

B sacchari wild type 938 62 010

189 prp UV mutant 436 564 090

189 Sucrosepropionatefeeding rates

920 180 127

Y3HVprp = 3HV yield from propionic acid

Maximum theoretical yield = 135 gg

P3HB-co-3HV from sucrose and propionic acid

3HB C4-monomer PHA

3HV ndash C5

Inactivation on the propionate

catabolic pathway

2MCC = 2 methyl citrate cycle

prpR

ORF1 ORF3 ORF4 ORF5ORF6 ORF7 ORF8 ORF9ORF2

prpB prpC acnM

EcoRIEcoRI EcoRIEcoRI

EcoRI

SalISalI

4 kbp 29 kbp 2 kbp 9 kbp

A

B

putative lipoprotein

52 bp

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

Characterization of mutants

a-1

a-2

a-3

b-1

b-2

c-1

c-2

d e

f

g

h

Random mutagenesis to identify genes related to biosynthetic pathway

KS AT P

P TEPKSi 10

2069 aa

KS AT DH ER KR P

P

PKSi 5

2585 aa

KS AT TEP

P

PKSi 11

2037 aa

KS AT P

P

P

P M H PPKSi 12

2637 aa

KS ATD

HM KR P

P C A RP

P

PKS-NRPS (PKSi6)

3981 aa

K

S

PKSIII

406 aa

PKSi 2KS AT M (ER)

K

RP

P

D

H

2262 aa

KS ATD

HM KR P

P

PKSi 1

2515 aa

KS AT DH MD

HKR P

P

PKSi 3

2576 aa

P

PKS ATD

HKR

PKSi 4

1777 aa

KS ATP

P

P

P TEPKSi 7

2145 aa

P

PKS AT DH M ER KRPKSi 8

2488 aa

P

PKS AT M RPKSi 9

2576 aa

K

S

A

T

K

R

D

H

E

R

T

E

M

P

P

H

P

R

Ketosynthase domain

Acyl transferase domain

Ketoreductase domain

Dehidrathese domain

Enoil reductase domain

Thioesterase domain

Methyltransferase domain

Acyl Carrier protrein domain

Hydrolase domain

Acyl CoA reductase domain

Ketosynthase domain

13 hypothetical PKS clusters identified in the E nigrum P16 genome Aminoacid numbers and domain positions are in scale

Epicolactone

New metabolite from Epicoccum nigrum

Associated to microbial control

Two PKS regulators were identified

Identification of the PKS gene cluster

Dr J Gregoacuterio C Gomez Dr Luiziana F Silva

PHA are accumulated as intracelular granules by bacteria

C4 amp C5

Short-chain length monomers

PHA scl

C6 amp C12

Medium-chain length

monomers PHA mcl

Naturally PHA-accumulating bacteria produce eitherPHAscl or PHAmcl in large amounts

Monomer composition is responsible for PHA properties and applications

730

Features influencing PHA monomer composition

Isolation amp evaluation of soil bacteria on the ability to produce PHB

from Sucrose Glucose and Fructose

Table 2 Production of P(3HB-co-3HV) from glucose plus propionic acid

Residual PHA

Strains CDWc

(gL)Carbohydrates

()

CDWc3HB

mol

3HV

molY3HVPrp

b

A eutrophus DSM 54 392 00 714 961 39 013

A latus DSM 1123 095 1018 146 550 450 007

Pcepacia DSM 50181 335 19 384 973 27 004

IPT-040 377 17 323 971 29 005

IPT-044 392 17 511 971 29 007

IPT-045 373 00 494 962 38 008

IPT-048 297 00 443 962 38 006

IPT-055 427 722 15 1000 00 000

IPT-056 360 313 309 985 15 002

IPT-076 506 19 568 971 29 010

IPT-083 489 52 568 969 31 010

IPT-086a 206 75 390 899 101 009

IPT-098 590 00 177 947 53 007

IPT-101 298 418 323 954 46 005

Gomez et al 1996

Improvement to

incorporate 3HV units

Strain Phenotype

strategy

3HB

mol

3HV mol

Y3HVprp

gg

B sacchari wild type 938 62 010

189 prp UV mutant 436 564 090

189 Sucrosepropionatefeeding rates

920 180 127

Y3HVprp = 3HV yield from propionic acid

Maximum theoretical yield = 135 gg

P3HB-co-3HV from sucrose and propionic acid

3HB C4-monomer PHA

3HV ndash C5

Inactivation on the propionate

catabolic pathway

2MCC = 2 methyl citrate cycle

prpR

ORF1 ORF3 ORF4 ORF5ORF6 ORF7 ORF8 ORF9ORF2

prpB prpC acnM

EcoRIEcoRI EcoRIEcoRI

EcoRI

SalISalI

4 kbp 29 kbp 2 kbp 9 kbp

A

B

putative lipoprotein

52 bp

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

KS AT P

P TEPKSi 10

2069 aa

KS AT DH ER KR P

P

PKSi 5

2585 aa

KS AT TEP

P

PKSi 11

2037 aa

KS AT P

P

P

P M H PPKSi 12

2637 aa

KS ATD

HM KR P

P C A RP

P

PKS-NRPS (PKSi6)

3981 aa

K

S

PKSIII

406 aa

PKSi 2KS AT M (ER)

K

RP

P

D

H

2262 aa

KS ATD

HM KR P

P

PKSi 1

2515 aa

KS AT DH MD

HKR P

P

PKSi 3

2576 aa

P

PKS ATD

HKR

PKSi 4

1777 aa

KS ATP

P

P

P TEPKSi 7

2145 aa

P

PKS AT DH M ER KRPKSi 8

2488 aa

P

PKS AT M RPKSi 9

2576 aa

K

S

A

T

K

R

D

H

E

R

T

E

M

P

P

H

P

R

Ketosynthase domain

Acyl transferase domain

Ketoreductase domain

Dehidrathese domain

Enoil reductase domain

Thioesterase domain

Methyltransferase domain

Acyl Carrier protrein domain

Hydrolase domain

Acyl CoA reductase domain

Ketosynthase domain

13 hypothetical PKS clusters identified in the E nigrum P16 genome Aminoacid numbers and domain positions are in scale

Epicolactone

New metabolite from Epicoccum nigrum

Associated to microbial control

Two PKS regulators were identified

Identification of the PKS gene cluster

Dr J Gregoacuterio C Gomez Dr Luiziana F Silva

PHA are accumulated as intracelular granules by bacteria

C4 amp C5

Short-chain length monomers

PHA scl

C6 amp C12

Medium-chain length

monomers PHA mcl

Naturally PHA-accumulating bacteria produce eitherPHAscl or PHAmcl in large amounts

Monomer composition is responsible for PHA properties and applications

730

Features influencing PHA monomer composition

Isolation amp evaluation of soil bacteria on the ability to produce PHB

from Sucrose Glucose and Fructose

Table 2 Production of P(3HB-co-3HV) from glucose plus propionic acid

Residual PHA

Strains CDWc

(gL)Carbohydrates

()

CDWc3HB

mol

3HV

molY3HVPrp

b

A eutrophus DSM 54 392 00 714 961 39 013

A latus DSM 1123 095 1018 146 550 450 007

Pcepacia DSM 50181 335 19 384 973 27 004

IPT-040 377 17 323 971 29 005

IPT-044 392 17 511 971 29 007

IPT-045 373 00 494 962 38 008

IPT-048 297 00 443 962 38 006

IPT-055 427 722 15 1000 00 000

IPT-056 360 313 309 985 15 002

IPT-076 506 19 568 971 29 010

IPT-083 489 52 568 969 31 010

IPT-086a 206 75 390 899 101 009

IPT-098 590 00 177 947 53 007

IPT-101 298 418 323 954 46 005

Gomez et al 1996

Improvement to

incorporate 3HV units

Strain Phenotype

strategy

3HB

mol

3HV mol

Y3HVprp

gg

B sacchari wild type 938 62 010

189 prp UV mutant 436 564 090

189 Sucrosepropionatefeeding rates

920 180 127

Y3HVprp = 3HV yield from propionic acid

Maximum theoretical yield = 135 gg

P3HB-co-3HV from sucrose and propionic acid

3HB C4-monomer PHA

3HV ndash C5

Inactivation on the propionate

catabolic pathway

2MCC = 2 methyl citrate cycle

prpR

ORF1 ORF3 ORF4 ORF5ORF6 ORF7 ORF8 ORF9ORF2

prpB prpC acnM

EcoRIEcoRI EcoRIEcoRI

EcoRI

SalISalI

4 kbp 29 kbp 2 kbp 9 kbp

A

B

putative lipoprotein

52 bp

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

Epicolactone

New metabolite from Epicoccum nigrum

Associated to microbial control

Two PKS regulators were identified

Identification of the PKS gene cluster

Dr J Gregoacuterio C Gomez Dr Luiziana F Silva

PHA are accumulated as intracelular granules by bacteria

C4 amp C5

Short-chain length monomers

PHA scl

C6 amp C12

Medium-chain length

monomers PHA mcl

Naturally PHA-accumulating bacteria produce eitherPHAscl or PHAmcl in large amounts

Monomer composition is responsible for PHA properties and applications

730

Features influencing PHA monomer composition

Isolation amp evaluation of soil bacteria on the ability to produce PHB

from Sucrose Glucose and Fructose

Table 2 Production of P(3HB-co-3HV) from glucose plus propionic acid

Residual PHA

Strains CDWc

(gL)Carbohydrates

()

CDWc3HB

mol

3HV

molY3HVPrp

b

A eutrophus DSM 54 392 00 714 961 39 013

A latus DSM 1123 095 1018 146 550 450 007

Pcepacia DSM 50181 335 19 384 973 27 004

IPT-040 377 17 323 971 29 005

IPT-044 392 17 511 971 29 007

IPT-045 373 00 494 962 38 008

IPT-048 297 00 443 962 38 006

IPT-055 427 722 15 1000 00 000

IPT-056 360 313 309 985 15 002

IPT-076 506 19 568 971 29 010

IPT-083 489 52 568 969 31 010

IPT-086a 206 75 390 899 101 009

IPT-098 590 00 177 947 53 007

IPT-101 298 418 323 954 46 005

Gomez et al 1996

Improvement to

incorporate 3HV units

Strain Phenotype

strategy

3HB

mol

3HV mol

Y3HVprp

gg

B sacchari wild type 938 62 010

189 prp UV mutant 436 564 090

189 Sucrosepropionatefeeding rates

920 180 127

Y3HVprp = 3HV yield from propionic acid

Maximum theoretical yield = 135 gg

P3HB-co-3HV from sucrose and propionic acid

3HB C4-monomer PHA

3HV ndash C5

Inactivation on the propionate

catabolic pathway

2MCC = 2 methyl citrate cycle

prpR

ORF1 ORF3 ORF4 ORF5ORF6 ORF7 ORF8 ORF9ORF2

prpB prpC acnM

EcoRIEcoRI EcoRIEcoRI

EcoRI

SalISalI

4 kbp 29 kbp 2 kbp 9 kbp

A

B

putative lipoprotein

52 bp

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

Dr J Gregoacuterio C Gomez Dr Luiziana F Silva

PHA are accumulated as intracelular granules by bacteria

C4 amp C5

Short-chain length monomers

PHA scl

C6 amp C12

Medium-chain length

monomers PHA mcl

Naturally PHA-accumulating bacteria produce eitherPHAscl or PHAmcl in large amounts

Monomer composition is responsible for PHA properties and applications

730

Features influencing PHA monomer composition

Isolation amp evaluation of soil bacteria on the ability to produce PHB

from Sucrose Glucose and Fructose

Table 2 Production of P(3HB-co-3HV) from glucose plus propionic acid

Residual PHA

Strains CDWc

(gL)Carbohydrates

()

CDWc3HB

mol

3HV

molY3HVPrp

b

A eutrophus DSM 54 392 00 714 961 39 013

A latus DSM 1123 095 1018 146 550 450 007

Pcepacia DSM 50181 335 19 384 973 27 004

IPT-040 377 17 323 971 29 005

IPT-044 392 17 511 971 29 007

IPT-045 373 00 494 962 38 008

IPT-048 297 00 443 962 38 006

IPT-055 427 722 15 1000 00 000

IPT-056 360 313 309 985 15 002

IPT-076 506 19 568 971 29 010

IPT-083 489 52 568 969 31 010

IPT-086a 206 75 390 899 101 009

IPT-098 590 00 177 947 53 007

IPT-101 298 418 323 954 46 005

Gomez et al 1996

Improvement to

incorporate 3HV units

Strain Phenotype

strategy

3HB

mol

3HV mol

Y3HVprp

gg

B sacchari wild type 938 62 010

189 prp UV mutant 436 564 090

189 Sucrosepropionatefeeding rates

920 180 127

Y3HVprp = 3HV yield from propionic acid

Maximum theoretical yield = 135 gg

P3HB-co-3HV from sucrose and propionic acid

3HB C4-monomer PHA

3HV ndash C5

Inactivation on the propionate

catabolic pathway

2MCC = 2 methyl citrate cycle

prpR

ORF1 ORF3 ORF4 ORF5ORF6 ORF7 ORF8 ORF9ORF2

prpB prpC acnM

EcoRIEcoRI EcoRIEcoRI

EcoRI

SalISalI

4 kbp 29 kbp 2 kbp 9 kbp

A

B

putative lipoprotein

52 bp

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

PHA are accumulated as intracelular granules by bacteria

C4 amp C5

Short-chain length monomers

PHA scl

C6 amp C12

Medium-chain length

monomers PHA mcl

Naturally PHA-accumulating bacteria produce eitherPHAscl or PHAmcl in large amounts

Monomer composition is responsible for PHA properties and applications

730

Features influencing PHA monomer composition

Isolation amp evaluation of soil bacteria on the ability to produce PHB

from Sucrose Glucose and Fructose

Table 2 Production of P(3HB-co-3HV) from glucose plus propionic acid

Residual PHA

Strains CDWc

(gL)Carbohydrates

()

CDWc3HB

mol

3HV

molY3HVPrp

b

A eutrophus DSM 54 392 00 714 961 39 013

A latus DSM 1123 095 1018 146 550 450 007

Pcepacia DSM 50181 335 19 384 973 27 004

IPT-040 377 17 323 971 29 005

IPT-044 392 17 511 971 29 007

IPT-045 373 00 494 962 38 008

IPT-048 297 00 443 962 38 006

IPT-055 427 722 15 1000 00 000

IPT-056 360 313 309 985 15 002

IPT-076 506 19 568 971 29 010

IPT-083 489 52 568 969 31 010

IPT-086a 206 75 390 899 101 009

IPT-098 590 00 177 947 53 007

IPT-101 298 418 323 954 46 005

Gomez et al 1996

Improvement to

incorporate 3HV units

Strain Phenotype

strategy

3HB

mol

3HV mol

Y3HVprp

gg

B sacchari wild type 938 62 010

189 prp UV mutant 436 564 090

189 Sucrosepropionatefeeding rates

920 180 127

Y3HVprp = 3HV yield from propionic acid

Maximum theoretical yield = 135 gg

P3HB-co-3HV from sucrose and propionic acid

3HB C4-monomer PHA

3HV ndash C5

Inactivation on the propionate

catabolic pathway

2MCC = 2 methyl citrate cycle

prpR

ORF1 ORF3 ORF4 ORF5ORF6 ORF7 ORF8 ORF9ORF2

prpB prpC acnM

EcoRIEcoRI EcoRIEcoRI

EcoRI

SalISalI

4 kbp 29 kbp 2 kbp 9 kbp

A

B

putative lipoprotein

52 bp

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

Monomer composition is responsible for PHA properties and applications

730

Features influencing PHA monomer composition

Isolation amp evaluation of soil bacteria on the ability to produce PHB

from Sucrose Glucose and Fructose

Table 2 Production of P(3HB-co-3HV) from glucose plus propionic acid

Residual PHA

Strains CDWc

(gL)Carbohydrates

()

CDWc3HB

mol

3HV

molY3HVPrp

b

A eutrophus DSM 54 392 00 714 961 39 013

A latus DSM 1123 095 1018 146 550 450 007

Pcepacia DSM 50181 335 19 384 973 27 004

IPT-040 377 17 323 971 29 005

IPT-044 392 17 511 971 29 007

IPT-045 373 00 494 962 38 008

IPT-048 297 00 443 962 38 006

IPT-055 427 722 15 1000 00 000

IPT-056 360 313 309 985 15 002

IPT-076 506 19 568 971 29 010

IPT-083 489 52 568 969 31 010

IPT-086a 206 75 390 899 101 009

IPT-098 590 00 177 947 53 007

IPT-101 298 418 323 954 46 005

Gomez et al 1996

Improvement to

incorporate 3HV units

Strain Phenotype

strategy

3HB

mol

3HV mol

Y3HVprp

gg

B sacchari wild type 938 62 010

189 prp UV mutant 436 564 090

189 Sucrosepropionatefeeding rates

920 180 127

Y3HVprp = 3HV yield from propionic acid

Maximum theoretical yield = 135 gg

P3HB-co-3HV from sucrose and propionic acid

3HB C4-monomer PHA

3HV ndash C5

Inactivation on the propionate

catabolic pathway

2MCC = 2 methyl citrate cycle

prpR

ORF1 ORF3 ORF4 ORF5ORF6 ORF7 ORF8 ORF9ORF2

prpB prpC acnM

EcoRIEcoRI EcoRIEcoRI

EcoRI

SalISalI

4 kbp 29 kbp 2 kbp 9 kbp

A

B

putative lipoprotein

52 bp

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

Features influencing PHA monomer composition

Isolation amp evaluation of soil bacteria on the ability to produce PHB

from Sucrose Glucose and Fructose

Table 2 Production of P(3HB-co-3HV) from glucose plus propionic acid

Residual PHA

Strains CDWc

(gL)Carbohydrates

()

CDWc3HB

mol

3HV

molY3HVPrp

b

A eutrophus DSM 54 392 00 714 961 39 013

A latus DSM 1123 095 1018 146 550 450 007

Pcepacia DSM 50181 335 19 384 973 27 004

IPT-040 377 17 323 971 29 005

IPT-044 392 17 511 971 29 007

IPT-045 373 00 494 962 38 008

IPT-048 297 00 443 962 38 006

IPT-055 427 722 15 1000 00 000

IPT-056 360 313 309 985 15 002

IPT-076 506 19 568 971 29 010

IPT-083 489 52 568 969 31 010

IPT-086a 206 75 390 899 101 009

IPT-098 590 00 177 947 53 007

IPT-101 298 418 323 954 46 005

Gomez et al 1996

Improvement to

incorporate 3HV units

Strain Phenotype

strategy

3HB

mol

3HV mol

Y3HVprp

gg

B sacchari wild type 938 62 010

189 prp UV mutant 436 564 090

189 Sucrosepropionatefeeding rates

920 180 127

Y3HVprp = 3HV yield from propionic acid

Maximum theoretical yield = 135 gg

P3HB-co-3HV from sucrose and propionic acid

3HB C4-monomer PHA

3HV ndash C5

Inactivation on the propionate

catabolic pathway

2MCC = 2 methyl citrate cycle

prpR

ORF1 ORF3 ORF4 ORF5ORF6 ORF7 ORF8 ORF9ORF2

prpB prpC acnM

EcoRIEcoRI EcoRIEcoRI

EcoRI

SalISalI

4 kbp 29 kbp 2 kbp 9 kbp

A

B

putative lipoprotein

52 bp

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

Isolation amp evaluation of soil bacteria on the ability to produce PHB

from Sucrose Glucose and Fructose

Table 2 Production of P(3HB-co-3HV) from glucose plus propionic acid

Residual PHA

Strains CDWc

(gL)Carbohydrates

()

CDWc3HB

mol

3HV

molY3HVPrp

b

A eutrophus DSM 54 392 00 714 961 39 013

A latus DSM 1123 095 1018 146 550 450 007

Pcepacia DSM 50181 335 19 384 973 27 004

IPT-040 377 17 323 971 29 005

IPT-044 392 17 511 971 29 007

IPT-045 373 00 494 962 38 008

IPT-048 297 00 443 962 38 006

IPT-055 427 722 15 1000 00 000

IPT-056 360 313 309 985 15 002

IPT-076 506 19 568 971 29 010

IPT-083 489 52 568 969 31 010

IPT-086a 206 75 390 899 101 009

IPT-098 590 00 177 947 53 007

IPT-101 298 418 323 954 46 005

Gomez et al 1996

Improvement to

incorporate 3HV units

Strain Phenotype

strategy

3HB

mol

3HV mol

Y3HVprp

gg

B sacchari wild type 938 62 010

189 prp UV mutant 436 564 090

189 Sucrosepropionatefeeding rates

920 180 127

Y3HVprp = 3HV yield from propionic acid

Maximum theoretical yield = 135 gg

P3HB-co-3HV from sucrose and propionic acid

3HB C4-monomer PHA

3HV ndash C5

Inactivation on the propionate

catabolic pathway

2MCC = 2 methyl citrate cycle

prpR

ORF1 ORF3 ORF4 ORF5ORF6 ORF7 ORF8 ORF9ORF2

prpB prpC acnM

EcoRIEcoRI EcoRIEcoRI

EcoRI

SalISalI

4 kbp 29 kbp 2 kbp 9 kbp

A

B

putative lipoprotein

52 bp

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

Table 2 Production of P(3HB-co-3HV) from glucose plus propionic acid

Residual PHA

Strains CDWc

(gL)Carbohydrates

()

CDWc3HB

mol

3HV

molY3HVPrp

b

A eutrophus DSM 54 392 00 714 961 39 013

A latus DSM 1123 095 1018 146 550 450 007

Pcepacia DSM 50181 335 19 384 973 27 004

IPT-040 377 17 323 971 29 005

IPT-044 392 17 511 971 29 007

IPT-045 373 00 494 962 38 008

IPT-048 297 00 443 962 38 006

IPT-055 427 722 15 1000 00 000

IPT-056 360 313 309 985 15 002

IPT-076 506 19 568 971 29 010

IPT-083 489 52 568 969 31 010

IPT-086a 206 75 390 899 101 009

IPT-098 590 00 177 947 53 007

IPT-101 298 418 323 954 46 005

Gomez et al 1996

Improvement to

incorporate 3HV units

Strain Phenotype

strategy

3HB

mol

3HV mol

Y3HVprp

gg

B sacchari wild type 938 62 010

189 prp UV mutant 436 564 090

189 Sucrosepropionatefeeding rates

920 180 127

Y3HVprp = 3HV yield from propionic acid

Maximum theoretical yield = 135 gg

P3HB-co-3HV from sucrose and propionic acid

3HB C4-monomer PHA

3HV ndash C5

Inactivation on the propionate

catabolic pathway

2MCC = 2 methyl citrate cycle

prpR

ORF1 ORF3 ORF4 ORF5ORF6 ORF7 ORF8 ORF9ORF2

prpB prpC acnM

EcoRIEcoRI EcoRIEcoRI

EcoRI

SalISalI

4 kbp 29 kbp 2 kbp 9 kbp

A

B

putative lipoprotein

52 bp

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

Improvement to

incorporate 3HV units

Strain Phenotype

strategy

3HB

mol

3HV mol

Y3HVprp

gg

B sacchari wild type 938 62 010

189 prp UV mutant 436 564 090

189 Sucrosepropionatefeeding rates

920 180 127

Y3HVprp = 3HV yield from propionic acid

Maximum theoretical yield = 135 gg

P3HB-co-3HV from sucrose and propionic acid

3HB C4-monomer PHA

3HV ndash C5

Inactivation on the propionate

catabolic pathway

2MCC = 2 methyl citrate cycle

prpR

ORF1 ORF3 ORF4 ORF5ORF6 ORF7 ORF8 ORF9ORF2

prpB prpC acnM

EcoRIEcoRI EcoRIEcoRI

EcoRI

SalISalI

4 kbp 29 kbp 2 kbp 9 kbp

A

B

putative lipoprotein

52 bp

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

Inactivation on the propionate

catabolic pathway

2MCC = 2 methyl citrate cycle

prpR

ORF1 ORF3 ORF4 ORF5ORF6 ORF7 ORF8 ORF9ORF2

prpB prpC acnM

EcoRIEcoRI EcoRIEcoRI

EcoRI

SalISalI

4 kbp 29 kbp 2 kbp 9 kbp

A

B

putative lipoprotein

52 bp

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

3HV content is dependent on propionic acid concentration

acnMprpC deletion

2MCC is more operative

at low prp concentrations

A second prp catabolic

pathway do exist

Control on 3HV content

Sucrosepropionic ratio in the feeding media

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

SucroseGlucoseXyloseGlycerolFatty acidsPlant oilsSoybean molassesOthers

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

Improving product yields inbiotechnological processes

bullKnockout or gene overexpression is noguarantee of success

bullA better knowledge of the metabolismis needed to engineer metabolism andimprove biotechnological processes

Vallino amp Stephanopoulos 1992

Metabolic engineering

advanced analytical tools to identify appropriate targets for genetic modifications

mathematical models to perform in silico design of optimized cell factoriesNielsen amp Jewett 2008 FEMS Yeast Res

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

Fluxes analysis is representative ofthe phenotype ofbiotechnologicalinterest

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

C (negligible)

dCdt = 0 (steady state)

Metabolic pathwayanalysis

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

Fluxes distribution for PHA productionby Pseudomonas sp from glucose

Optimal fluxes distribution for PHA productionby Pseudomonas sp from glucose

PHAmcl

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

049

048

001

165

PHA production by wild type B sacchari Fluxes under the theoretical

maximum global PHA yield

049

001

173

124116

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

Brazilian biodiversity as newplatforms

GlucoseXyloseGlycerolFatty acidsPlant oilsSoybeanmolasses

Pseudomonas spB sacchariE coliPlatforms for

PHARhamonolipids13-PropanediolOthers

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical

NWO

MES-Cuba

TUDelft

JGregoacuterio C GomezLuiziana F SilvaMarilda Keico TaciroKarel Olavarria GamezRogeacuterio S GomesJohana K Bocanegra-RodriguezThatiane T MendonccedilaGabriela C LozanoLinda P Guaman BautistaLiege A KawaiLucas Garbini CespedesDiana Carolina Tusso PizonBernardo Ferreira CamiloKelli Lopes RodriguesCesar W Guzman MorenoRafael NahatCarlos Thandara Garcia RavelliJuliano CherixEdmar Ramos de Oliveira FilhoRuben Sanchez (UENF)Aline Carolina C LemosAlexandre A AlvesAelson L Santos

Karen L AlmeidaOdalys Rodriguez GamezArelis Abalos RodriguezJhoanne Hansen

Amanda B Flora

Galo A C Le Roux - EPUSPCarlos A M RiascosPaulo AlexandrinoAndreacute Fujita IME USPJuliana Cardinali Rezende

Andreas K Gombert (UNICAMP)Walter M van GulikAljoscha WahlReza Maleki SeifarJ J (Sef) HeijnenKirsten Steinbusch ndash Waste2ChemicalNiels van Stralen ndash Waste2Chemical