#ppl 0g ctusbdut · vii national conference bifi2015 zaragoza, february 4-6, 2015 book of...

Book Of Abstracts

4-6 February 2015, Edificio I+D, Campus Río Ebro

Contents

I Invited contributions 1

II BIFI research lines 11

III Short communications 21

IV Posters 47

i

VII National Conference BIFI2015Zaragoza, February 4-6, 2015

BOOK OF ABSTRACTS

ii

Part I

Invited contributions

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Plant root release of phenolics and flavins upon Fe deficiency

A. M. Alvarez Fernandez, P. Siso-Terraz, Y. Gogorcena, A. Abadıa

and J. Abadıa

Department of Plant Nutrition, Estacion Experimental de Aula Dei (CSIC)

Plant roots release many chemical compounds into the growth medium. Released com-

pounds can play roles in different plant processes, including below-ground plant defense as

well as nutrient acquisition. Secreted phytochemicals can solubilize and/or mobilize soil nu-

trients that are poorly available to plants. The components of plant root exudates are many,

complex and dependent on the specific plant species and growth environments, and have been

poorly characterized so far. Iron (Fe) deficiency is one of the most limiting factors for crop

production and induces chlorosis in about 30 percent of crops worldwide. Crops affected

include those grown in calcareous, high pH soils that cause the precipitation of Fe in spar-

ingly soluble ferric oxy-hydroxydes forms. In order to overcome the Fe solubility problem,

many bacteria, fungi and plants commonly rely on the synthesis and secretion of specialized

chemicals to facilitate Fe mining from the soil. For instance, it is well known that Gramineae

export multidentate amino acid ferric chelators, belonging to the mugineic acids family, to

acquire unavailable Fe from the soil. Many non-Gramineae plant species/cultivars tolerant to

Fe deficiency are also able to accumulate in roots and export to the rhizosphere a plethora

of compounds such as phenolics and flavins in response to low Fe availability (Cesco et al.,

2010). However, the identity of these compounds, the mechanisms of transport from the

roots to the rizhosphere, as well as their role in Fe acquisition are still poorly known. The

presentation will discuss the characterization of such exudates, both in model plant species

(Arabidopsis thaliana and Medicago truncatula) and in agronomically relevant crop species

(Solanum lycopersicum and Prunus persica), using high-resolution mass spectrometry instru-

mentation and new data processing techniques. Also, the talk will give an overview of the

most recent findings on the molecular components involved in the synthesis and transport of

these secreted compounds in model plants, which support that the root secretion of phenolics

and flavins is an integral part of the Fe uptake machinery. Finally, the studies on how this

root exports improve the plants’ ability to cope with limited Fe availability will be examined.

[1]Cesco S., Neumann G., Tomasi N., Pinton R., and Weisskopf L. (2010) Plant Soil, 329,

1-25

Acknowledgments: Supported by the Spanish Ministry of Economy and Competitiveness

(grants AGL2012-31988 and AGL2013-42175-R, co-financed with FEDER) and the Aragon

Government (groups A03 and A44). Patricia Siso-Terraza was supported by a MINECO-FPI

contract.

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Granulysin as a new anti-tumoral treatment, preclinical phase

A. Anel, S. Al-Wasaby, A. Aporta, D. de Miguel, J. Naval, J. Pardo, L.

Martınez-Lostao and B. Conde

Department of Biochemistry and Molecular and Cell Biology,

University of Zaragoza (SPAIN)

Granulysin is a protein present in the granules of human CTL and NK cells, with cytolytic

activity against microbes and tumors. Previous work for our group characterized the mecha-

nism by which granulysin induced apoptosis in vitro on hematological tumor cells. Mrore re-

cent work demonstrated that granulysin was cytotoxic against cells from chronic lymphocytic

leukemia patients but not on normal human peripheral blood lymphocytes. We have now stu-

dided the in vivo effect of recombinant granulysin on tumor development in an athymic “nude”

mice model bearing human mammary MDA-MB-231 or multiple myeloma H929–derived

xenografts. Granulysin prevented the in vivo development of detectable MDAMB- 231-derived

tumors. In addition, recombinant granulysin, independently from its LPS content, was able

to completely eradicate H929-derived tumors. All granulysin-treated tumors exhibited signs

of apoptosis induction and an increased NK cell infiltration inside the tumor tissue comparing

to control ones. No deleterious effects of recombinant granulysin doses used in this study were

observed on the dermis or on the internal organs of the animals. This work opens the door

to the initiation of preclinical and possibly clinical studies for the use of granulysin as a new

anti-tumoral treatment.

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Structural and Local-Spectroscopic Studies on

(Bio)-Nanomaterials by TEM

Raul Arenal1,2

1 Laboratorio Microscopıas Avanzadas, INA,

University of Zaragoza, 50018 Zaragoza(SPAIN)2 Fundacion ARAID, 50018 Zaragoza(SPAIN)

In the last two decades, transmission electron microscopes (TEM) have undergone a

large number of improvements allowing 100 meV (even few tens of meV) energy reso-

lutions for a close to one angstrom electron beam and low acceleration voltages. These

performances offer new possibilities for probing the atomic configuration, the optical,

dielectric and electronic properties of nanomaterials with unprecedented spatial infor-

mation, as well as for studying the atomic configuration of nanostructures. Today, I

will present a selection of the recent works involving all these matters. These works will

concern the study of the atomic structure & configuration of nanostructures (including

doped carbon nanotubes and bio-nanomaterials) as well as the opto-electronic prop-

erties studies carried out via electron energy loss spectroscopy (EELS) measurements

of different kind of nano- objects. These works will illustrate the excellent capabil-

ities offered by the use of a Cs probe corrected STEM, combined with the use of a

monochromator, to study these properties within a very good spatial resolution.

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How to deploy infrastructures on cloud and not become a

sys-admin

Ignacio Blanquer

GRyCAP, Universidad Politenica de Valencia, Spain

Cloud computing has become extremely popular in the last years. The ability of tempo-

rary acquiring a large infrastructure without upfront costs is appealing for many use cases,

including scientific research. However, the proper usage, deployment, configuration and mon-

itoring of virtualized cloud infrastructures is cumbersome and requires some degree of system

administration skills.

The Grid and High Performance (GRyCAP - www.grycap.upv.es) research group of the

UPV has developed an ecosystem of tools and services to assist on this point. This ecosystem

provides solutions for managing rich metadata from Virtual Machine Images, using on-the-fly

configuration and configuration based on reusable and platform-independent recipes, man-

aging elasticity from batch queues workload, reducing unnecessary energy consumption on

physical back-ends and reducing the fragmentation of VMs.

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Neutral evolution and the acceleration of the molecular clock

Jose A. Cuesta1,2

1 GISC, Universidad Carlos III de Madrid 28911 Leganes, Madrid (SPAIN)2 BIFI, Universidad de Zaragoza, 50018 Zaragoza (SPAIN)

Large sets of genotypes give rise to the same phenotype because phenotypic expression

is highly redundant. Accordingly, a population can accept mutations without altering

its phenotype, as long as the genotype mutates into another one on the same set. By

linking every pair of genotypes that are mutually accessible through mutation, geno-

types organize themselves into neutral networks (NN). These networks are known to be

heterogeneous and assortative, and these properties affect the evolutionary dynamics of

the population. By studying the dynamics of populations on NN with arbitrary topol-

ogy we analyze the effect of assortativity, of NN (phenotype) fitness, and of network

size. We find that the probability that the population leaves the network is smaller

the longer the time spent on it. This progressive ”phenotypic entrapment” entails a

systematic increase in the overdispersion of the process with time and an acceleration

in the fixation rate of neutral mutations.

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Genetics and genomics of Mycobacterium tuberculosis, from

the bench to the clinic.

Jesus Gonzalo-Asensio1 and Carlos Martın2

1 BIFI-CIBERES, Universidad de Zaragoza2 IIS Aragon-CIBERES, Universidad de Zaragoza

Epidemic diseases such as tuberculosis, AIDS and malaria are poverty-related and

represent a scourge for developing countries. Patient surveillance, antibiotic treatment

and prophylactic vaccination are successful preventive measures to control epidemic

diseases. However, these measures are not easily applied to Mycobacterium tuberculosis

due to the high emergence of drug resistant strains and the variable efficacy of the

present vaccine BCG. With this scenario, the development of a new vaccine represents

the best cost-effective tool to achieve tuberculosis eradication in the future.

In this presentation, we summarize 14 years of basic research on Mycobacterial genet-

ics and its application to tuberculosis control. Specifically, we detail the step-by-step ap-

proach to develop the live-attenuated M. tuberculosis vaccine candidate MTBVAC from

the initial discovery of the phoP virulence gene by the University of Zaragoza/Institut

Pasteur in 2000 to the final vaccine manufacture by BIOFABRI in 2010 and the first

human clinical trial in 2014. We present a review of the key scientific events leading

to vaccine characterization, which include i) the identification of the PhoP regulatory

network, ii) dissection of the attenuation-associated phenotypes, iii) protective efficacy

experiments in animal models and iv) immunogenicity and apoptosis elicited by the

vaccine candidate.

Finally, we outline our current basic research derived from the knowledge of the

vaccine candidate including the study of a small RNA regulated by PhoP that might

impact on the immunogenicity of the strain. We also report our future plan to develop

multivalent vaccines by secreting heterologous antigens in MTBVAC.

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PELE: a Monte Carlo technique for protein engineering and

drug design

Vıctor Guallar

BSC-Centro Nacional de Supercomputacion, Barcelona, Spain

Understanding the protein-ligand/substrate interaction mechanism requires the de-

scription of ligand migration and the binding site induced fit. Such study involves

dynamic time scales on the range of micro-seconds, a non-trivial task for current com-

putational methods. Despite these difficulties, it has centered a great deal of effort

from the molecular dynamics community, with its highest point, probably, in the recent

design and development of the ANTON machine [1].

Here we will present an alternative to MD techniques. Using technological advances

in protein structure prediction, we have recently introduced PELE (protein energy land-

scape explorations). PELE combines a Monte Carlo stochastic approach with protein

structure prediction algorithms and is capable of accurately reproducing long time scale

processes in only few hours of CPU (typically no more than an overnight computing

period) [2,3]. In this talk will summarize the technique, and some of our latest studies

including both enzyme engineering and protein-ligand binding mechanistic studies for

drug design.

[1] Shan, Y.; Kim, E. T.; Eastwood, M. P.; Dror, R. O.; Seeliger, M. A.; Shaw, D. E.

Journal of the American Chemical Society 2011, 133, 9181.

[2] Cossins, B. P.; Hosseini, A.; Guallar, V. Journal of Chemical Theory and Compu-

tation 2012.

[3] Borrelli, K. W.; Vitalis, A.; Alcantara, R.; Guallar, V. J. Chem. Theory Comput.

2005, 1, 1304.

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Opening data at Gobierno de Aragon

Jose Marıa Subero

Direccion General de Nuevas Tecnologıas del Gobierno de Aragon

50071 Zaragoza

Opening public data is a path that governments must walk. They started this way

through the open government initiatives but nowadays there is a lot of work to do yet.

At the Aragon Government, we are trying to improve data access to the public with

the vision of having a default publication of the information.

To walk this path it is necessary to take some key decisions related with technology,

legality, user services, data management and so on. During the 7th BIFI National

Conference we will try to answer and discuss about some of the solutions taken to

these key decisions. Furthermore, we will present some of the most innovative services

created by Aragon Open Data and we will draft some of the future ideas about what

is going to happen in the open data world.

10

Part II

BIFI research lines

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New (and ot so new) drugs against tubercolosis

J.A. Ainsa1,2,3,4, C. Villellas2,3,4, R. Bailo2,3,4, A. Lucia2,3,4, S.

Ramon-Garcıa2,3,4 and B. Gracia2,3,4

1 BIFI, University of Zaragoza, Spain2 Departamento de Microbiologıa, Medicina Preventiva y Salud Publica,

Facultad de Medicina, Universidad de Zaragoza3 Instituto de Investigacion Sanitaria Aragon ( IIS-Aragon), Spain.4 CIBER Enfermedades Respiratorias, Instituto de Salud Carlos III,

Spain.

The use of antimicrobials is one of the most effective ways to combat infectious dis-

eases. In the case of tuberculosis, the scarce number of effective drugs available (just

five first-line drugs, of which at least four are used in the standard treatment of tubercu-

losis caused by drug susceptible strains), and the increasing incidence of drug resistant

tuberculosis leave very limited therapeutic options. This situation has prompted in-

vestigation in new drugs, new targets and new therapies, and at the same time some

already existing drugs are being modified, repurposed or included in new formula-

tions with the hope of developing new medicines for curing tuberculosis.We have been

studying efflux pumps as a mechanism of intrinsic drug resistance in Mycobacterium

tuberculosis and demonstrated that avoiding efflux is a key feature for a successful new

drug, hence reinforcing the interest in developing efflux inhibitors. By combining ge-

netic and microbiological techniques, several other potential drug candidates are being

explored.

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Modelizacion de biomoleculas

Pierpaolo Bruscolini1,2 and Fernando Falo1,3

1 BIFI, Universidad de Zaragoza, Zaragoza (SPAIN)2 Departamento de Fısica Teorica, Universidad de Zaragoza, Zaragoza(SPAIN)3 Departamento de Fısica de la Materia Condensada,

Universidad de Zaragoza, Zaragoza(SPAIN)

Se presentan las lineas de investigacion y resultados mas relevantes obtenidos en los

dos ultimos anos en el campo de la modelizacion de biomoleculas

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Protein Intrinsic Disorder in Cellular Function and Disease

Nunilo Cremades Casasin1,2

1 BIFI, University of Zaragoza, Spain2 Departamento de Bioquımica y Biologıa Molecular,

Universidad de Zaragoza, Spain

The sequence-structure-function paradigm that states that the amino acid sequence

of a protein determines its definite ordered 3D structure, which in turn is required

for its biological function, has been challenged by the discovery of many proteins and

protein domains that posses specific functions whilst lacking stable ordered structures

under physiological conditions. Such intrinsically disordered proteins (IDPs) and pro-

tein regions (IDPRs) are involved in cellular functions such as regulation and signaling,

although a few cases of enzymes devoid of rigid ordered structure have also been re-

cently reported, further defying long-established assumptions on the structure-function

and the lock- and-key paradigms. In addition, numerous IDPs are associated with

human diseases, including cancer and neurodegenerative disorders.

During my talk, I will review some of our recent studies on structural disorder in-

volved in protein function [1], particularly in catalytic function [2], as well as in disease,

specifically in amyloid conditions, for which either disorder-to-order [3,4] or order-to-

disorder-to-order [5] transitions are required.

[1] Aprile et al. PLoS One. 2013 Jun 28;8(6):e67961. doi: 10.1371/ journal.pone.0067961.

Print 2013.

[2] Zambelli and Cremades et al. 2012. Mol Biosyst. Jan;8(1):220-8.

[3] Cremades et al. 2012. Cell. May 25;149(5):1048-59.

[4] Chen et al. submitted

[5] Dhulesia et al. 2010. J Am Chem Soc. Nov 10;132(44):15580-8.

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OXPHOS system functional genetics

Patricio Fernandez-Silva 1,2

1 BIFI, University of Zaragoza, Spain2 Departamento de Bioquımica y Biologıa Molecular,

Universidad de Zaragoza, Spain

GENOXPHOS group focus is the molecular and genetic study of the mitochondrial

oxidative phosphorylation (OXPHOS) system. Our reserach tries to cover from the ba-

sic regulatory mechanisms of this system, its biogenesis and structural organization up

to the understanding, at the molecular level, of the diseases caused by its dysfunction

and the assay of potential therapeutic strategies. Some of the recent achievements and

contributions of our group, including the proposal of a new respiratory chain organi-

zation model (Science, 2013, 340(6140):1567-70. doi: 10.1126/science.1230381) will be

presented.

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ETP4HPC, the European technology Platform for High

Performance Computing

Guillermo Losilla

BIFI, Universidad de Zaragoza, Zaragoza(SPAIN)

BIFI has recently joined ETP4HPC, the European Technology Platform for High-

Performance Computing (http://www.etp4hpc.eu). It is an strategic, industry-led fo-

rum which provides a framework for stakeholders (technology providers, supercomput-

ing centers, researchers, ISVs...) to define European HPC technology research priorities

and action plans. In this talk we will introduce the project, its main objectives (in-

cluding the Strategic Research Agenda for HPC in Europe and the HPC contractual

Public-Private Partnership recently signed with the European Comission) and the ben-

efits BIFI could expect to obtain by joining this initiative.

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The Physics of Human Systems

Yamir Moreno1,2,3

1 BIFI, University of Zaragoza, Spain2 Department of Theoretical Physics, University of Zaragoza, Spain3 Complex Networks and Systems Lagrange Lab,

Institute for Scientific Interchange, Turin, Italy

Physics has been extremely successful in describing our natural world, from the very

small to the very large scales. In particular, statistical physicists are used to deal

with many particles and this is perhaps the reason why large scale social systems have

attracted so much attention within the physics community. However, as we will argue in

this talk, the study of human collective behavior is not as easy as dealing, for instance,

with ideal gases. The reasons are multiple, among which one can mention the fact that

we do not know what are the laws describing most human behaviors and that in many

dynamical processes details really matter. Through the dissemblance of 3 different

examples of human collective behavior, we will show what are the experimental and

theoretical challenges in the study of large scale social systems and propose a way to

tackle such problems.

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Disecting allosteric interactions in proteins

Adrian Velazquez-Campoy1,2

1 BIFI, University of Zaragoza, Spain2 Fundacion ARAID, Zaragoza Spain

Allosteric interactions constitute the energetic and structural molecular basis for bio-

logical function regulation and control. Allosterism involves the interplay and coupling

between ligand binding events and conformational changes in biological targets, result-

ing in cooperative phenomena. A brief review of allosterism and binding cooperativity

in proteins will be given, and the methodology for studying allosteric biological systems

by isothermal titration calorimetry will be described.

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20

Part III

Short communications

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Collecting and analyzing social data

Alberto Alcolea


The social data, data published on Internet on social networks, message boards and

other social sources, are ambiguous, unstructured, and they are not formatted according

to any ontology. The purpose of this talk is to expose two projects developed on the

Institute for Biocomputation and Physics of Complex Systems (BIFI) and Kampal (a

spin-off of BIFI) about the collection and the analysis of social data: Aragon Open

SocialData and Kampal Social.

The first one, Aragon Open SocialData, is a project whose aim is to collect all social

information generated in Aragon or related with it and give to third people a simple

API to reuse all data collected in researches, applications and reports.

Kampal Social provides a listening system whose goal is to study and measure the

social impact on Internet of events, campaigns and other happenings through the col-

lection and the analysis of social data.

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Development of a platform for the creation of participatory

experiments of sociological nature

Julio Alberto Arenere


The goal is to explain how to develop a web-based platform of surveys that allow users

easily, creation, modification and publication of surveys and obtaining the results of

this. Besides to the modification of this platform for the development of the experiment

”¿Como son nuestros voluntarios?”

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.Identification of compounds that inhibit NUPR1 protein, a

key therapeutic target in Pancreatic Ductal Adenocarcinoma

(PDAC)

M. Arruebo1, A. Lanas2, J. L. Neira3,4, J. L. Iovanna5, A.

Velazquez-Campoy4,6 and O. Abian1,4

1 IIS Aragon / Instituto Aragones de Ciencias de la Salud-IACS,

Zaragoza, Spain2 IIS Aragon / Servicio de Digestivo - Hospital C.U. Lozano Blesa,

Zaragoza, Spain3 Instituto de Biologıa Molecular y Celular, Universidad Miguel Hernandez,

Elche, Spain4 BIFI, Universidad de Zaragoza, Spain5 Centre de Recherche en Cancerologie de Marseille (CRCM),

INSERM U1068, Marsella, France

PDAC, the most common type of pancreatic cancer, is one of the most aggressive human

cancers and worse prognosis because of its remarkable capacity for invasion and metastasis.

PDAC exhibits an extremely low survival rate (< 3 − 4% at 5 years), due to its difficult

diagnosis in early stages, and its high resistance to chemotherapy and radiotherapy. Hence,

there is an urgent need to develop new and effective therapies against this disease.

Protein NUPR1 (Nuclear Protein 1), also known as p8, is a nuclear protein over-expressed in

several types of human cancers (breast, thyroid, skin...), especially in PDAC, where NUPR1 is

essential for the development and progression of the tumor. It has been demonstrated in mice

studies that, in this kind of tumors, after blocking NUPR1 expression in pancreatic cancer

cells, the formation of tumors can be prevented, confirming its key role in PDAC.

NUPR1 protein is a small (82 aminoacid residues), basic and multifunctional protein which

belongs to the group of proteins called intrinsically unstructured or intrinsically disordered

proteins (IDPs). Its interaction with its different partners promotes the mutual stabilization

of their structures in a particular conformation and the formation of fuzzy, but biologically

active complex. NUPR1 fuzzy complexes are involved in the regulation of important cellular

processes, such as transcription of genes, cellular cycle, or apoptosis.

In our group, we have developed a new strategy for identifying compounds that stabilize

the native structure of p8 and block NUPR1 intracellular interactions. The in vitro selected

compounds inhibited NUPR1 interactions with one of its cellular partners and efficiently

blocked NUPR1 function in cell-based studies.

References:

“p8 and Prothymosin Alpha” Cedric Malicet, Jean Charles Dagorn, Jose Luis Neira, Juan

L. Iovanna.* [Cell Cycle 5:8, 829-830, 15 April 2006];

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“Nuclear protein 1 promotes pancreatic cancer development and protects cells from stress

by inhibiting apoptosis” Tewfik Hamidi et.al.J Clin Invest. 2012;122(6): 2092–2103.

doi:10.1172/JCI60144.

“Emerging role of nuclear protein 1 (NUPR1) in cancer biology” Uttio Roy Chowdhury &

Rajeev S. Samant & Oystein Fodstad & Lalita A. Shevde. Cancer Metastasis Rev (2009)

28:225–232 DOI 10.1007/s10555-009-9183-x

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.Mechanical unfolding of DNA quadruplexes: a microscopic

view of the role of the central ionsA. E. Bergues-Pupo1,2,J. R. Arias-Gonzalez3, M. C. Moron4, A.

Fiasconaro5 and F. Falo1,2

1 BIFI, Universidad de Zaragoza, 50018 Zaragoza, Spain.2 Departamento de Fısica de la Materia Condensada,

Facultad de Ciencias, Universidad de Zaragoza3 ICMA, Universidad de Zaragoza, 50009 Zaragoza, Spain.4 IMDEA Nanocienciam, Madrid, Spain5 School of Mathematical Sciences, Queen Mary University of London, UK

DNA quadruplexes are non canonical structures in which guanine rich sequences can self-

assembly in the presence of monovalent cations to form a four-stranded structure. The basic

unit of such structures is the quartet, a planar arrangement of guanines held by Hoogsteen

hydrogen bonding [1]. Quadruplex have been observed in vivo at the telomere and in gene

promoter regions [2,3]. They seem to play a major structural and regulatory role in the

chromosomal maintenance and in the controlling of gene expression. Quadruplexes can adopt

manifold topologies characterized by their relative strand orientation. Specifically, human

quadruplex telemore can be folded in the so called parallel and antiparallel conformations [4].

AFM and optical tweezers have been used to study the mechanical stability of these different

quadruplex topologies [5-7]. The results of such kind of experiments show that antiparallel

conformations are mechanically more stable than the parallel ones [5,6]. These mechanical

differences may provide insights in the variety of flexibility, ligand binding affinity and folding

pathway of the quadruplexes.

In this work we use molecular dynamic simulations to study the mechanical unfolding of

different short segments of DNA quadruplexes. We look at the role of the central ions in the

unfolding of a parallel and an antiparallel conformations. The overall mechanical stability of

the quadruplexes is found to be sensitive to the coordination of the ions and their neighbor

guanines.

[1] Burge S. , Parkinson G.N., Hazel P., Todd A.K., and Neidle S. (2006) Nucleic Acids

Res. 34, 5402.

[2] Lam E.Y., Beraldi D., Tannahill D., and Balasubramanian S. (2013) Nat. Commun. 4,

1796.

[3] Siddiqui-Jain A., Grand C.L., Bearss D.J., and Hurley L.H. (2002) Proc. Natl. Acad.

Sci. U.S.A. 99, 11593.

[4] Parkinson G.N., Lee M.P., and Neidle S., (2002) Nature 417, 876.

[5] Yu Z., Schonhoft J.D., Dhakal S., Bajracharya R., Hegde R., Basu S., and Mao H. (2009)

J. Am. Chem. Soc. 131, 1876.

[6] Messieres M.; Chang J.Ch.; Brawn-Cinani B.; La Porta A. (2012) Phys. Rev. Lett. 109,

058101.

[7] Garavıs M.; Bocanegra R.; Herrero-Galan E.; Gonzalez C.; Villasante A.; Arias-Gonzalez

J.R. (2013) Chem. Commun. 49, 6397.

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Ultrafast single electron spin manipulation in 2D quantum

dots with optimally controlled time-dependent electric fields

Jorge Budagosky 1 and Alberto Castro1,2

1 BIFI, Universidad de Zaragoza, Zaragoza(SPAIN)2 Fundacion ARAID, Zaragoza (SPAIN)

We have studied theoretically the possibility of ultra-fast manipulation of a single

electron spin in 2D semiconductor quantum dots, by means of high-frequency time-

dependent electric fields. The electron spin degree of freedom is excited through spin-

orbit coupling, and the procedure may be enhanced by the presence of a static magnetic

field. We use quantum optimal control theory to tailor the temporal profile of the

electric field in order to achieve the most effective manipulation. The scheme predicts

significant control over spin operations in times of the order of picoseconds –an ultrafast

time scale that permits to avoid the effects of decoherence if this scheme is to be used

as a tool for quantum information processing.

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FurB, the zinc regulator in Anabaena sp. PCC 7120, plays a

key role in the cyanobacterial oxidative stress response

Violeta C. Sein-Echaluce1,2 and M. Luisa Peleato 1,2

1 Departamento de Bioquımica y Biologıa Molecular y Celular,

Universidad de Zaragoza, Spain2 BIFI, Universidad de Zaragoza, Spain

Iron and zinc are essential nutrients whose homeostasis is tightly controlled by members of

the Fur superfamily in the cyanobacterium Anabaena sp. PCC7120. Iron levels inside the cells

are tightly controlled by the FurA paralogue, whereas FurB/Zur is responsible for maintaining

zinc levels [1]. The link between iron metabolism and oxidative stress management is well

documented, however, little is known about the connection between zinc homeostasis and the

oxidative stress response in cyanobacteria.

Previous studies in our laboratory have shown that, when overexpressed in textitEscherichia

coli, FurB improved cell survival against oxidative stress. FurB also protected DNA against

hydroxyl radical damage in vitro [2]. Therefore, in order to investigate the possible cor-

relation between FurB and the oxidative stress response in Anabaena, furB deletion and

furB-overexpressing strains have been constructed and the consequences of FurB imbalance

evaluated.

The results show that the lack of FurB increased sensitivity to H2O2, whereas an excess

of the protein enhanced oxidative stress resistance of the cyanobacterium. Both mutants ex-

hibited an altered expression of different genes coding for proteins involved in the oxidative

stress response, namely SodA, catalases and several peroxiredoxins. Transcriptional and bio-

chemical analyses unveiled that the appropriate level of FurB is required for proper control of

the oxidative stress response and allowed us to identify major antioxidant enzymes as novel

members of the FurB/Zur regulon [3].

[1] Napolitano M., et al. (2012). Journal of Bacteriology 194(10): 2426-2436.

[2] Lopez-Gomollon S., et al. (2009). Biochemical Journal 418(1): 201.

[3] Sein-Echaluce V.C., et al (2014). Environmental Microbiology. doi: 10.1111/1462-

2920.12628.

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Conversion of 5-hydroxymethylfurfural to furandicarboxylic

acid in an enzymatic reaction cascade

Juan Carro Aramburu1, P. Ferreira2,3, L. Rodrıguez1, A. Prieto1, A.

Serrano1, B. Balcells1, A. Arda1, J. Jimenez-Barbero1, A. Gutierrez3,

R. Ullrich4, M. Hofrichter4 and A. T. Martınez1

1 Centro de Investigaciones Biologicas-CSIC, Madrid, Spain2 Departamento de Bioquımica y Biologıa Molecular y Celular,

Universidad de Zaragoza, Spain3 BIFI, Universidad de Zaragoza, Spain4Department of Bio- and Environmental Sciences,

International Instute of Zittau, Germany

Oxidative conversion of 5-hydroxymethylfurfural (HMF) is of biotechnological interest for

the production of renewable (lignocellulose based) platform chemicals, such as 2,5-furandicar-

boxylic acid (FDCA). The ability of fungal aryl-alcohol oxidase (AAO) to oxidize HMF is re-

ported here for the first time, resulting in a nearly complete conversion into 2,5-formylfurancar-

boxylic acid (FFCA) in a few hours. The reaction starts by alcohol oxidation yielding 2,5-

diformylfuran (DFF) that is rapidly converted into FFCA by carbonyl oxidation, most prob-

ably without leaving the enzyme active site. This agrees with the similar catalytic efficiency

of the enzyme oxidizing HMF and DFF and its very low activity on 2,5-hydroxymethylfuran-

carboxylic acid (that could not be detected by GC-MS). However, AAO was found to be

unable to directly oxidize the carbonyl group in FFCA, and only modest amounts of FDCA

are formed from HMF (most probably by chemical oxidation of FFCA by the H2O2 previously

generated by AAO). Since aldehyde oxidation by AAO proceeds via the corresponding geminal

diols (aldehyde hydrates), the different carbonyl oxidation rates could be related to the low

hydration degree of FFCA compared with DFF. The conversion of HMF was completed by

introducing a fungal unspecific heme peroxygenase that uses the H2O2 generated by AAO to

transform FFCA into FDCA, albeit more slowly than the previous AAO reactions. By adding

this peroxygenase when FFCA production by AAO has been completed, transformation of

HMF into FDCA can be attained in a reaction cascade where O2 is the only cosubstrate

needed, and water the only by-product formed.

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Rescuing compound bioactivity in a secondary cell-based

screening by using gamma cyclodextrins as a molecular carrier

R. Claverıa1,4,5, A. Velazquez-Campoy1,2,3 and O. Abian1,4,5

1 BIFI, Universidad de Zaragoza, Spain2 Department of Biochemistry and Molecular and Cell Biology,

University of Zaragoza, Zaragoza, Spain.3 Fundacion ARAID, Zaragoza 50018, Spain4 IACS-IIS Aragon, Zaragoza, Spain;

Centro de Investigacion Biomedica en Red en el Area Tematica de

Enfermedades Hepaticas y Digestivas (CIBERehd), Spain.5 Instituto de Investigacion Sanitaria de Aragon (IIS Aragon).

Although specific inhibitors have been approved for Hepatitis C Virus (HCV) clinical ther-

apy very recently, resistance-associated mutations have already been reported for those drugs,

compromising their long-term efficacy. Therefore, there is an urgent need for new anti-HCV

agents with low susceptibility to resistance-associated mutations. NS3 protease protein has

been considered a target for drug development since its identification two decades ago. Sev-

eral inhibitors of this enzyme have been identified. Three of them exhibited good efficacy

in vitro but they had poor efficacy in cellular assays. In order to enhance the cell uptaking

of these compounds, they were complexed with +-cyclodextrin and afterwards, their efficacy

was tested using a replicon system that mimic the virus cycle in cells. Two of them increased

5 and 16 times their efficacy in comparison when they were offered as free compounds. But

the most significant result came with one compound that did not show any antiviral activity

in cells when it was free but its +-cyclodextrin complexed exhibited an efficacy value con-

centration of 5 µM . So, the antiviral efficacy of these compounds can be improved or even

rescued using +-cyclodextrin as carrier molecule.

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BOOK OF ABSTRACTS

.

Structural organization in multiplex networks

Emanuele Cozzo1 and Yamir Moreno1,2,3



Given a multiplex network M , it has naturally associated two networks of reduced dimen-

sionality: the aggregated network and the network of layers [1]. We define the reduced form

R(M) of the original multiplex as the Cartesianproduct of the aggregated network and the

network of layers. We associate to both M and R(M) a density matrix ρ(M) and ρ(R(M))

respectively, given by the combinatorial Laplacian normalized by the degree sum.We consider

the relative entropy of ρ(M) with respect to ρ(R(M)) . At a graph-theoretic level this mea-

sure can be understood as the amount of regularities and nontrivial symme- tries introduced

or destroyed by the aggregation operation. In this sense, it can be interpreted as a measure of

the complexity of the original multiplex network. Capitalizing on the interlacing results in [1],

we are able to give some bounds for the relative entropy. Besides, we show that, depending

on the weight of the inter-layer coupling the multiplex undergoes structural transition.

[1] R. Sanchez-Garcia, E. Cozzo, and Y. Moreno, “Dimensionality reduction and spectral

properties of multilayer networks”, Physical Review E 89, 052815 (2014).

32


BOOK OF ABSTRACTS

.

Molecular Interaction Model Between the Human

Apoptosis-inducing Factor (hAIF) and the Nuclease

Cyclophilin A (CypA)

E. Fernandez-Alonso1, I. Yruela2,3, M. Medina3,4 and P. Ferreira3,4

1 Servicio de Farmacia, Hospital C. U. Lozano Blesa, Zaragoza2 Estacion Experimental de Aula Dei (EEAD-CSIC), Zaragoza3 BIFI, Universidad de Zaragoza, Spain4 Departamento de Bioquımica y Biologıa Molecular y Celular,

Universidad de Zaragoza

The Apoptosis-Inducing Factor (hAIF) is a mitochondrial flavoenzyme involved in pro-

grammed cell death. After a cellular apoptotic insult, hAIF is released from the mitochondria

and translocated to the nucleus where it induces apoptosis. However, the hAIF lethal activity

depends on the histone H2AX and CypA, probably through their interactions in a ternary com-

plex named ”DNA-degradosome” [1]. In healthy cells, AIF presents a NADH-oxidoreductase

activity that plays an essential but unknown role in mitochondrial energy production. Since

the hAIF redox state influences its conformation, and by extension its pro-apoptotic activity,

has been postulated that both functions are related [2].

In this work, the interaction between human AIF (hAIF) and CypA has been analyzed

using bioinformatic tools. The results show that the hAIF -CypA interaction depends on the

hAIF redox state. In the hAIF oxidized state three interaction regions are proposed, two of

them, T141-G156 and W196-Y211, also present in its reduced state and a third, T521- S532,

only observed in the oxidized state. However, in the hAIF reduced state two new regions

appear, G466-L486 and I581-E596, as a result of conformational changes that occur upon

NADH binding and protein reduction. These conformational changes create a cavity that

enables access of CypA, facilitating its binding to hAIF.

[1] Artus C., Boujrad H., Bouharrour A., Brunelle M.N., Hoos S., Yuste V.J., Lenormand

P., Rousselle J.C., Namane A., England P., Lorenzo H.K., Susin S. A. (2010) Embo J, 29,

1585-99.

[2] Ferreira P., Villanueva R., Martınez-Julvez M., Herguedas B., Marcuello C., Fernandez-

Silva P., Hermoso J.A., Lostao A., Susin SA. and Medina M. (2014) Biochemistry, 53, 4204-

15.

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BOOK OF ABSTRACTS

.

Isolation and Characterization of Hemopexin as an acute

Phase protein in Pigs

N. Garcıa, L. Soler, M. A. Alava and F. Lampreave

Departamento de Bioquımica y Biologıa Molecular y Celular,


Acute phase proteins (APPs) are serum proteins whose concentrations change during pe-

riods of infection, inflammation and trauma. Although the role of APPs is not well known,

it is thought APPs have an important role against infections and repairing damaged tissues.

Hemopexin (Hx) has been described as a positive APP in many species including human, rat

and mouse. In this work, we have achieved the isolation of Hx from pig sera by a proce-

dure comprising a precipitation with amonium sulphate followed by Cibacron Blue-Sepharose

Cromatography. Hx has been used to immunize rabbits for the preparation of a specific

antiserum. The response of this protein was studied in pigs under different acute phase

experimental models: - Pancreas transplantation surgery (n=11) (PTS); - Experimental in-

flammation after turpine injection (n=3) (TI); - Actinobacillus Pneumoniae infection (n=3)

(API).

A decrease in the Hx concentration was observed in sera from PTS pigs. The Hx levels

decrease from 1, 70±0, 34 mg/ml before the surgery (range: 1,18- 2,20) to 1, 22±0, 44 mg/ml,

24 hours after (range: 0,57-1,81). Moderate decreases were observed in TI processes, being

25% less than the initial concentration. After API, Hx concentration % exhibited a reduction

of about 30% initial values. Thus, Hx seems likely to act as a negative APP in pigs.

More work is needed to evaluate the usefulness of these findings to understand the biochem-

ical applications of this protein in the context of the acute phase processes.

34


BOOK OF ABSTRACTS

.

Accounting of HPC resources at BIFI

Arturo Giner


2014 has been an outstanding year for BIFI’s HPC resources; the record of CPU hours

delivered by our supercomputers (Memento/Caesaraugusta & Terminus) has been surpassed.

Thus, during the past year, a total of 21,14 millions of CPU hours have been accounted in

our main HPC systems, some of them reaching an utilization level > 75%. The principal con-

sumers of this vast computing time have been BIFI’s researchers (and their collaborators) and

external users through the Spanish Supercomputing Network, the Hosted Clusters program

and billed services. Through this talk, we will explore the accounting data gathered during the

last year (active users, hours consumed, applications used, level of parallelization...), trying

to obtain a profile of the average user of our HPC resources.

35


BOOK OF ABSTRACTS

.

Reputation drives cooperative behavior and network

formation in human groups

C. Gracia-Lazaro1 J. Cuesta1,2, A. Ferrer1, Y. Moreno1,3,4 and A.

Sanchez1,2

1 BIFI, University of Zaragoza, Spain2 GISC, Universidad Carlos III de Madrid, Spain3 Department of Theoretical Physics, University of Zaragoza, Spain4 Complex Networks and Systems Lagrange Lab,


Cooperativeness is a defining feature of human nature. Theoreticians have suggested several

mechanisms to explain this ubiquitous phenomenon, including reciprocity, reputation, and

punishment, but the problem is still unsolved. Here we show, through experiments conducted

with groups of people playing an iterated Prisoner’s Dilemma on a dynamic network, that it is

reputation what really fosters cooperation. While this mechanism has already been observed

in unstructured populations, we find that it acts equally when interactions are given by a

network that players can reconfigure dynamically. Furthermore, our observations reveal that

memory also drives the network formation process, and cooperators assort more, with longer

link lifetimes, the longer the past actions record. Our analysis demonstrates, for the first

time, that reputation can be very well quantified as a weighted mean of the fractions of past

cooperative acts and the last action performed. This finding has potential applications in

collaborative systems and e-commerce.

36


BOOK OF ABSTRACTS

.

Ehrenfest statistical dynamics in chemistry: study of

decoherence effects

J. L. Alonso1,2,3, P. Bruscolini1,2,3, A. Castro1,4, J.

Clemente-Gallardo1,2,3, J. C. Cuchı4 and J. A. Jover-Galtier1,2

1 BIFI, Universidad de Zaragoza, Zaragoza(SPAIN)2 Departamento de Fısica Teorica, Unviersidad de Zaragoza,

Zaragoza(SPAIN)3 Unidad asociada IQFR-BIFI4 Fundacion ARAID, Zaragoza (SPAIN)5 Departament d’Enginyeria Agroforestal, ETSEA-Universitat de Lleida,

Lleida (SPAIN)

Our work proves that the method of Statistical Ehrenfest Dynamics introduced in [1 and

2] to approach nonadiabaticity is able to predict, in a rigorous ab initio way, changes in

the purity of the quantum state of the electrons of a molecule. As an example we have

studied the dynamical properties of ionized sodium molecules. Our formalism is based on a

statistical description of the mixed quantum-classical system, which can be marginalized to

define distributions for the nuclei only, or a density matrix which encodes the quantum state

of the electronic subsystem. Nonetheless, the evolution is defined as a combined statistical

system. Our method provides thus a procedure to model molecular dynamics in a simple way,

without implementing algorithmically the decoherence effects or adding ad-hoc terms to the

dynamics. The evolution of the molecular density matrix in our formalism takes into account,

in a natural way, the decoherent term introduced in the decay of mixing formalism [3 and 4].

This not only supports, dynamically, that formalism but also encourages the use of the SED

for non-adiabatic processes, provided that you do not perform experiments in which quantum

effects of the nuclei are directly observed.

[1] Alonso J. L., Castro A., Clemente-Gallardo J., Cuchı J. C., Echenique P., and Falceto

F. (2011) J. Phys. A: Math. Theo. 44, 395004.

[2] Alonso J. L., Clemente-Gallardo J., Cuchı J. C., Echenique P., and Falceto F. (2012) J.

Chem. Phys. 137, 054106.

[3] Zhu C., Jasper A. W., and Truhlar D. G. (2005) J. Chem. Theo. Comp. 1, 527.

[4] Truhlar D. G. (2007) in Quantum Dynamics of Complex Molecular Systems (edited by

D. A. Micha and I. Burghardt), pp. 227–243, Springer, Berlin, Germany.

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.

Application of Finite-Temperature String Method to the

Study of Hydride Transfer Between Anabaena FNR and

NADP+/H

I. Lans1,M. Medina2,3, E. Costa3, G. Hummer4, M. Garcıa-Viloca5, J. M.

Lluch5 and A. Gonzalez-Lafont5

1 Laboratory of Molecular Neuropharmacology and Bioinformatics,

Institut de Neurociencies and Unitat de Bioestadıstica,

Universitat Autonoma de Barcelona, 08193 Bellaterra, Spain.2 BIFI, Universidad de Zaragoza, Spain3 Department of Biochemistry and Molecular and Cell Biology,

University of Zaragoza, Zaragoza, Spain.4 Department of Chemistry, King’s College London,

London SE1 1UL, United Kingdom.5 Departament de Quımica and Institut de Biotecnologia i de Biomedicina,

Universitat Autonoma de Barcelona, 08193 Bellaterra (Barcelona), Spain.

Ferredoxin-NADP+ reductase (FNR) catalyses the electron transfer from ferredoxin (Fd) to

NADP+ to produce reducing power as NADPH during the photosynthesis. The final hydride

transfer event between FNR and the nucleotide is a reversible process. Two different tran-

sient charge-transfer complexes form prior to and upon hydride transfer, FNRrd-NADP+ and

FNRox-NADPH, and regardless of the hydride transfer direction. Experimental structures

for the FNRox:NADP+ interaction obtained under different conditions suggested a series of

conformational rearrangements that might contribute to attain the catalytically competent

complex, but so far no experimental information is available about the structure of this com-

plex. In this study a structure, close to the transient catalytically competent interaction of

Anabaena FNR with its coenzyme, NADP+, obtained from a molecular dynamics approach

has been initially used to undertake a fully microscopical simulation of the hydride transfer

processes between Anabaena FNRrd/FNRox and NADP+/H with the corresponding solvation

shell of water molecules. A dual level QM/MM hybrid approach has been used to describe the

potential energy surface of the whole system. MD calculations using the finite-temperature

string method with the WHAM method have provided the potential of mean force (PMF) of

the hydride transfer processes. The results confirm that the structural model of the reactants

evolves to a catalytic competent transition state through very similar free energy barriers for

the forward and reverse reactions, in good agreement with the hydride transfer experimental

rate constants reported in this system.

[1] Lans I, Medina M, Rosta E, Hummer G, Garcia-Viloca M, Lluch JM, and Gonzalez-

Lafont A. (2012). J Am Chem Soc.134,20544-53

38


BOOK OF ABSTRACTS

.

Contact-based social contagion in multiplex networks

S. Meloni1 E. Cozzo1, R. Alvarez1 and Y. Moreno1,2,3



Given a multiplex network M , it has naturally associated two networks of reduced dimen-

sionality: the aggregated network and the network of layers [1]. We define the reduced form

R(M) of the original multiplex as the Cartesianproduct of the aggregated network and the

network of layers. We associate to both M and R(M) a density matrix ρ(M) and ρ(R(M))

respectively, given by the combinatorial Laplacian normalized by the degree sum.We consider

the relative entropy of ρ(M) with respect to ρ(R(M)) . At a graph-theoretic level this mea-

sure can be understood as the amount of regularities and nontrivial symme- tries introduced

or destroyed by the aggregation operation. In this sense, it can be interpreted as a measure of

the complexity of the original multiplex network. Capitalizing on the interlacing results in [1],

we are able to give some bounds for the relative entropy. Besides, we show that, depending

on the weight of the inter-layer coupling the multiplex undergoes structural transition.

39


BOOK OF ABSTRACTS

.

7 non-stop running year Janus supercomputer overview

Sergio Perez-Gaviro1,2

1 BIFI, Universidad de Zaragoza, Zaragoza(SPAIN)2 Fundacion ARAID, Zaragoza (SPAIN)

The Special Purpose Computer Janus [1,2] was designed and developed as a multipur-

pose reprogramable supercomputer 7 years ago. Janus is based on a Field-Programmable-

Gate-Array (FPGA) architecture, which permits researchers reprogram the computer’s

hardware connections structure. During this time, Janus has been focused on the

study and simulation of spin glasses, paradigm of complex systems. Indeed it has

revealed as a very fruitful scientific computer for the study of these disordered sys-

tems. We were able to reach simulation times up to 0.1 seconds of an experiment

( 1011 Monte Carlo sweeps) for very large lattices and low temperatures. Several

scientific works performed thanks to Janus have been published in high impact sci-

entific journals, as in the Proceedings of the National Academy of Science or in the

Physical Review Letters. Moreover, recently our last work [3] deserved its inclusion

in 2014 December month’s ’Highlights’ section of the Journal Statistical Mechanics

(http://iopscience.iop.org/1742-5468/page/Highlights) Encouraged by the good results,

the Janus Collaboration decided to go an step further developing and designing the new

generation Janus dedicated computer, named JanusII. More information in our web site:

http://www.janus-computer.com/ During this presentation I will introduce the reasons

why a special purpose computer is needed for studying such a systems. I will also give a

brief overview through the Janus dedicated machine generation, from Janus to JanusII,

and I will briefly discuss some of the successful aims achieved with Janus during its life.

Note: Janus is now open to other researchers not belonging to the Janus Collabora-

tion. Any application, as molecular dynamics, digital signal processing, cryptography

or speech recognition among others, which can be fitted in a fpga architecture, will be

very welcome!

[1] Belletti F. et al. (2008) Computer Physics Communications 178 (3), p.208-216.

[2] Belletti F. et al. (2009) Computing in Science & Engineering 11-1, 48-58.

[3] Baity-Jesi M. et al. (2014) J. Stat. Mech. P05014.

[4] Baity-Jesi M. et al. (2014) Computer Physics Communications 185, 550-559

(2014).

40


BOOK OF ABSTRACTS

.

Revisiting the riboflavin kinase catalytic cycle of bacterial

FAD Synthetase.

Marıa Sebastian1,2 and Milagros Medina1,2

1 BIFI, Universidad de Zaragoza, Spain2 Departamento de Bioquımica y Biologıa Molecular y Celular,


The FAD Synthetase (FADS) from Corynebacterium ammoniagenes is a bifunctional en-

zyme that catalyses the transformation of riboflavin (RF) into flavin adenine dinucleotide

(FAD) in two steps.[1] First RF is phosphorilated to flavin mononucleotide (FMN) by a

riboflavin kinase activity, being subsequent converted into FAD by an adenylyl-transferase

activity. These activities are located in two different modules of the enzyme: the C-terminal

domain performs the riboflavin kinase activity (RFK), while the N-terminal module is respon-

sible from the FMN transformation into FAD. The FADS from the non pathogenic organism

C. ammoniagenes shows structural similarities with those from other pathogens such as M.

tuberculosis or S. pneumonia, being enzymes from C. ammoniagenes ususally used as models

for other organisms. Due to the essentiality of the catalytic activities of FADS for cell survival

and to their structural differences related to mammal enzymes involved in similar functions,

FADS appears as a potential drug target.[2] In this context, a deeper characterization of its

catalytic cycle is advantageous in the development of new antibacterial drugs. In this work,

we use the rapid mixing technique, stopped flow, to identify and quantify the different indi-

vidual steps of the RFK activity catalytic cycle. This is, to determine the ligands binding

and dissociation order, and the characteristic constants that defin every single process. [3]

Due to the complexity of the FADS catalytic cycle, we used a truncated form of the enzyme

consisting on the fully active RFK domain.

[1] Frago, S., et al., Structural analysis of FAD synthetase from Corynebacterium ammoni-

agenes. BMC Microbiol, 2008. 8: p. 160.

[2] Serrano, A., et al., The prokaryotic FAD Synthetase family: a potential drug target.

Current Pharm Design, 2013. In press.

[3] Frago, S., A. Velazquez-Campoy, and M. Medina, The puzzle of ligand binding to

Corynebacterium ammoniagenes FAD synthetase. J Biol Chem, 2009. 284(11): p. 6610-

9.

41


BOOK OF ABSTRACTS

.

Characterization of acute phase proteins associated to HDL

particles in canine and porcine sera

L. Soler1, N. Garcıa1, P. D. Eckersall2, M. A. Alava1 and F. Lampreave1


Universidad de Zaragoza2 Institute of Biodiversity, Animal Health and Comparative Medicine,

University of Glasgow, Bearsden Rd, Glasgow, G61 1QH, Scotland, UK

Veterinary sciences are essentially aimed at understanding biological mechanism related to

the animal health and welfare. In one hand dogs are sociable animals considered the main

company animals as pet, on the other hand pork is the most consumed meat from terrestrial

animals. Achievements in pig production and dog welfare it is related with one increase in an-

imal health. Acute phase proteins (APPS) are serum proteins that change their concentration

after infection, inflammation or trauma and this change could be useful to disease diagnosis or

for monitoring health through their use as biomarkers. It has been showed that in these situ-

ations lipoprotein metabolism is greatly modified. In recent studies a high number of human

plasmatic proteins associated with HDL particles were found. Following this approach our

aim was to find APPs associated to lipid fractions in canine and porcine species. Firstly, sera

were fractionated by gel filtration chromatography which allowed us to separate LDL, HDL

and other protein fractions in several collected fractions. Secondly, lipid-associated proteins

were purified by affinity binding chromatography using calcium hydrate silicate allowing us to

concentrate several proteins that in whole serum are in low concentration. In this comparative

study, analysis of the main APPs was performed by Western Blotting and MALDI-TOF. In

both species they showed the same affinity pattern. ITIH4, Apo A-I and C-reactive protein

were found associated to lipid fraction, and specially ITIH4 and Apo A-I in HDL fraction.

Results also showed that, as is known, Pig-MAP (major acute phase protein in pigs also called

ITIH4 in other species) and Apo A-I (negative acute phase protein) show the most change in

their concentration in acute phase response. Apo A-I loss in HDL during acute phase response

is related with pro-inflammatory activity associated to HDL particles. Other proteins studied

as haptoglobin, α1-acid glicoprotein and albumin were not found to be associated to lipids.

42


BOOK OF ABSTRACTS

.Free Energy Characterization of the Mechanical Dissociation

of Biological Complexes from Force Spectroscopy Experiments

R. Tapia-Rojo1,2, C. Marcuello3,4, C. Gomez-Moreno1,4, A. Lostao3,4, J.

J. Mazo2,5 and F. Falo1,2

1 BIFI, Universidad de Zaragoza, 50018 Zaragoza, Spain.2 Departamento de Fısica de la Materia Condensada,

Facultad de Ciencias, Universidad de Zaragoza3 INA, Universidad de Zaragoza, 50018 Zaragoza, Spain.4 Departamento de Bioquımica y Biologıa Molecular y Celular,

Universidad de Zaragoza, 50009 Zaragoza, Spain5 ICMA, Universidad de Zaragoza, 50009 Zaragoza, Spain.

Free energy landscape analysis has become a key tool fo understanding the physical behavior

of a large array of biomolecules, including conformational changes in macromolecules, nucleic-

acid and protein folding, or binding affinity of complexes. Since the arrival of single-molecule

techniques [1], big interest has been set onto the influence of mechanical perturbations onto

biomolecules, with outstanding studies about mechanical unfolding of proteins, or mechanical

unbinding of biological complexes, just to name a few. Here, we make a theoretical and ex-

perimental study of the mechanical unbinding of two protein-protein complexes, Fd-FNR and

Fd-FNR. In order to explore this process, we propose a model which assumes that the free

energy profile can be characterized by two magnitudes, a free energy barrier controlling the

kinetic behavior, and the dissociation free energy, governing the equilibrium properties. By

performing numerical simulations on the model, we are able to recover the free energy barrier

height and the dissociation free energy with the help of dynamic force spectroscopy theory [2]

and Jarzysnki equality [3], respectively. In addition, we analyze dynamic force spectroscopy

AFM experiments on the two proposed protein-protein complexes, yielding biologically rele-

vant values for both magnitudes, where, remarkably, the equilibrium free energies obtained

from Jarzynski equality match those obtained from calorimetry bulk experiments [4]. Our

work follows thus a twofold aim. On the one hand, we propose a general model for under-

standing mechanical dissociation of biological complexes, helping in gaining insight about the

physics of the process. Second, we put forward a robust analysis protocol for dynamic force

spectroscopy experiments which allows for a full characterization of the free energy profile

which governs this process, obtaining both kinetic and equilibrium magnitudes.

[1] Ritort. F, (2006) J. Phys.: Condens. Matter 18, R531-R583

[2] Dudko O. K., Hummer G., and Szabo A. (2006) Phys. Rev. Lett. 96, 108101

[3] Jarzynski C. (1997) Phys. Rev. Lett. 78, 2690

[4] Martınez-Julvez M. Medina M. and Velazquez-Campoy A. (2009) Biophys. J. 96,

4966–4975

43


BOOK OF ABSTRACTS

.

Towards a rational design of Saccharomyces cerevisiae Gas2

glycomimetics

J. Valero-Gonzalez1, I. Delso2, T. Tejero2 and R. Huertado-Guerrero1

1 Instituto de Biocomputacion y Fisica de Sistemas Complejos (BIFI),

Universidad de Zaragoza, 50018 Zaragoza, Aragon, Spain.2 Departamento de Sıntesis y Estructura de Biomoleculas,

Instituto de Sıntesis Quımica y Catalisis Homogenea (ISQCH),

Universidad de Zaragoza, CSIC, 50009 Zaragoza, Aragon, Spain.

Glycosyl hydrolases and transglycosylases play a determinating role in fungal cell wall dy-

namics. Among these enzymes, the transglycosylases family remodeling the β-1,3 glucan

have shown to be essential in yeast and in Asp. fumigatus. Therefore, these enzymes are

highly interesting as therapeutic targets [1]. To tackle a rationale design of glycomimetics for

this family, we chose the Saccharomyces cerevisiae Gas2 (ScGas2) for practical purposes: its

structure had been solved and it was very close to the essential Gel4 from Asp. Fumigatus

[2]. Even though ScGas2 shows the typical glycosyl hydrolase folding, its activity presents a

balance among hydrolysis and transglycosylation depending on the length of the substrate.

For example, ScGas2 is inactive against laminaripentaose but it is a pure hydrolase against

laminarihexaose and heptaose and becomes a transglycosylase against larger-sized laminari-

oligosaccharides. In the case of Gel family, laminarinonaose was the minimal tested size to

have transglycosylase activity [3]. Here, a combination of docking studies, synthesis of gly-

comimetics, fluorescence spectroscopy and STD-NMR experiments reveal that a minimum

of three glucose units linked via a β-(1,3) linkage and a hydrophobic radical are required to

achieve molecular recognition at the binding donor and acceptor sites in ScGas2, respectively.

Overall, our data set up the basis for the design of more potent glycomimetics that could be

useful to inhibit essential enzymes such as Gel4.

[1] Latge, J. P. (2007) Mol. Microbiol. 66 279–290

[2] Hurtado-Guerrero, R., Schuttelkopf, A. W., Mouyna, I., Ibrahim, A. F. M., Shepherd,

S., Fontaine, T., van Aalten, D. M. F. (2009).The Journal of Biological Chemistry, 284(13),

8461–8469

[3] Hartland R. P., Fontaine T., Debeaupuis J. P., Simenel C., Delepierre M., Latge J. P.

(1996) J. Biol. Chem, 271, 26843

44


BOOK OF ABSTRACTS

.

Cloud computing resources and projects at BIFI

R. Valles, J. Ibar and C. Gimeno


Cloud technology has evolved a lot specially in the recent years. It allows the usage of

virtualized computing, storage, network resources in a simple and effective way. The resources

are very flexible and thanks to virtualization technologies different operating systems like

Windows and Linux can be executed equally. For this reasons, among others, cloud computing

is growing in adoption by companies and researchers. Cloud computing resources and the

different projects in which BIFI is participating will be outlined in this presentation.

45


BOOK OF ABSTRACTS

46

Part IV

Posters

47


BOOK OF ABSTRACTS

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Identificacion y cuantificacion de receptores de melatonina en

el tracto reproductor de morueco.

D. Aguilar-Recarte, A.E. Gaspar-Torrubia, M Gonzalez-Arto, R

Perez-Pe, T Muino-Blanco, A Casao-Gascon and J.A.Cebrian-Perez

Department of Biochemistry and Molecular and Cellular Biology,

IUCA, Faculty of Veterinary, University of Zaragoza, Spain

La hormona melatonina es una de las sustancias biologicamente activas mas antiguas de los

organismos vivos y desempena una gran variedad de funciones. Una gran parte de estas fun-

ciones estan mediadas por un tipo de GPCRs (G-Protein Coupled Receptors) que intervienen

en rutas de regulacion activando diferentes vıas de transduccion de senal. La melatonina es

sintetizada principalmente por la glandula pineal durante la noche y, recientemente, estudios

llevados a cabo por nuestro grupo de investigacion, a partir de la determinacion de la presencia

constante de melatonina en plasma seminal ovino, han demostrado la existencia de enzimas

de sıntesis en el tracto reproductor de morueco.

De este modo, podrıa existir una regulacion propia en el tracto reproductor del macho,

independiente de los ritmos circadianos, que serıa responsable de los niveles de melatonina en

su interior, y estos niveles podrıan estar controlados por receptores especıficos de la hormona

en sus diferentes tejidos.

Con el fin de dilucidar la existencia de esta posible regulacion del tracto genital masculino,

se realizaron una serie de estudios en los distintos tejidos del aparato reproductor (testıculo,

epidıdimo y glandulas accesorias) centrados en demostrar la existencia de los dos receptores

principales: MT1 y MT2, es decir, receptores de melatonina 1 y 2. Mediante RT-PCR se

demostro la expresion genica de dichos receptores en los tejidos estudiados, mientras que

la q-PCR permitio cuantificar las diferencias de esta expresion. Por otra parte, mediante

Western-Blot se demostro la traduccion y presencia de estos receptores en la mayorıa de

los tejidos en mayor o menor medida, lo que implica la existencia activa de un proceso de

regulacion de los niveles de melatonina en el tracto reproductor de morueco.

La relevancia de dichos resultados radica en la existencia de los dos receptores de melatonina

en el tracto reproductor y, por consiguiente, en la actuacion de diferentes vıas de regulacion

asociadas a estos, que regulan la concentracion de la hormona en el tracto y su presencia en

plasma seminal, factor importante a la hora de determinar la supervivencia y maduracion de

celulas espermaticas, lo cual posee un papel decisivo en la capacidad fecundante y en otros

parametros reproductivos.

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BOOK OF ABSTRACTS

.

Purification and partial characterization of the recombinant

Fur protein from the plant pathogen Erwinia amylovora

N. Blasco1,2, L. Botello-Morte1,2,M. Fillat1,2, T. Bes 1,2 and M. L.

Peleato1,2


Universidad de Zaragoza2 BIFI, Universidad de Zaragoza, Spain

Fur (ferric uptake regulation) protein belongs to a DNA-binding protein family that regu-

lates iron responsive genes1. Erwinia amylovora is a plant pathogen, causing the fire blight

disease on pear, apple, and other rosaceous plants. The ability to infect of the pathogen is

closely related to iron metabolism2 and for this reason, the fur gene from the pathogen E.

amylovora CFBP 1430 have been cloned and over expressed successfully in E. coli BL21 (DE3).

Recombinant Fur from the pathogen has been purified using a Cu2+ IMAC affinity column

without His-tag, due to the His-rich sequence present in its structure. The experimental data

show that Fur oligomerizes in solution, with possible involvement of disulphide bridges. The

protein contains structural zinc but only present in the reduced form. The results obtained

could be helpful in producing protein with defined oligomerization states, and the obtained

high yield of the protein will permit further characterization.

Acknowledgements: we thank Dr Miguel Cambra and Dr Ana Palacio for their kind cession

of E. amylovora CFBP 1430 DNA.

[1] M. F. Fillat, (2014) The FUR (ferric uptake regulator) superfamily: Diversity and

versatility of key transcriptional regulators. Arch Biochem Biophys. (2014) 546:41-52.

[2] T. H. M. Smits, B. Duffy, (2011) Genomics of iron acquisition in the plant pathogen

Erwinia amylovora: insights in the biosynthetic pathway of the siderophore desferrioxamine

E. Arch Microbiol (2011) 193:693–699.

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BOOK OF ABSTRACTS

.

Analysis of the interactions involving the Anabaena Ferric

uptake regulator (Fur) paralogs using bacterial two hybrid

assays

N. Blasco1,2, M. Fillat1,2, T. Bes 1,2, M. L. Peleato1,2 and

L. Botello-Morte1,2



Fur (ferric uptake regulator) proteins constitute a large family of transcriptional regula-

tors present in most prokaryotes, exhibiting a wide diversity of functions. The filamentous,

nitrogen-fixing cyanobacterium Anabaena PCC 7120 contains three Fur paralogs. FurA is a

global regulator acting as a hub, which connects iron homeostasis, oxidative stress defense and

other relevant metabolic pathways. FurB controls zinc homeostasis and it is a key factor in the

oxidative stress response in Anabaena. Finally, little is known about the third paralog FurC,

which has been recently proposed as the peroxide-stress regulator in this cyanobacterium.

Previous results from our laboratory suggested a regulatory role for FurC over the DNA

binding activity of FurA and FurB, although it is unable to bind furA or furB promoters.

The presence of FurC resulted in an enhanced interaction of FurA with DNA, in contrast to

the inhibitory effect produced over the FurB-DNA binding. To get more insights into the

putative interactions of the three Fur paralogs present in Anabaena, fur genes were cloned

into the pKT25, pKNT25, pUT18 and pUT18C plasmids to perform bacterial two hybrid

assays (BACTH). The formation of heterodimers, as well as homodimers of every protein,

was analyzed in detail.

BACTH assays demonstrated that Fur proteins were able to homodimerize in vivo. Al-

though this phenomenon is a common feature among transcriptional regulators, no previous

evidence of in vivo homodimerization of any Fur protein has been reported in the litera-

ture. Moreover, the formation of heterodimers of Anabaena Fur paralogs has also been shown

by this technique. This result is in good agreement with the suggested modulation of the

transcriptional activity of both FurA and FurB regulators exerted by FurC.

51


BOOK OF ABSTRACTS

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Structural study on stabilization of the complex formed by

human Apoptosis Inducing Factor (hAIF) and its coenzyme

NAD+

J. E. Casaos1, R. Villanueva1, P. Ferreira1,2, M. Medina1,2 and M.

Martınez-Julvez1,2



Apoptosis-Inducing Factor (AIF) is a mitochondrial-flavoprotein involved in an alterna-

tive caspase-independent cell death mechanism. After a stimulus of programmed cellular

death (PCD) and the action of proteases, its pro-apoptotic form (hAIF∆1−102) is released

to the intermembrane space, translocated to the nucleus where induces typical chromatolysis

of independent caspases death. Deficiency in this enzyme has been associated with several

mitochondrial pathologies. Studies on mouse enzyme (mAIF), together with those on hu-

man enzyme (hAIF) produced in our group, have demonstrated that upon reduction of the

flavin cofactor of AIF by NADH, conformational changes lead to AIF dimerization. The

formed hAIF∆1−102rd:2 NAD+ complex is stabilized by interface interactions involving E413,

N424, E426, R430, S431 and R449 residues and presents two binding sites for NAD+.The

E413A/R422A/R430A mutant is unable to dimerize and has different redox characteristics

compared to the WT as well as different pro-apoptotic behavior. Here, we report the crystal-

lographic structure of the E413A/R422A/R430AhAIF∆1−102ox specie to analyze the relevance

of the dimer formation in the NADH reductase activity of hAIF. Its structure has been com-

pared with that of the WT enzyme, in oxidized and reduced state, and overall both fold

similarly. However, the 532-534 residues, located in a long and flexible region (509-559) that

apparently occludes access to the redox active site, show different conformations and the num-

ber of stabilizing interactions at the dimer interface is reduced in the case of the mutant. In

addition, E413A/R422A/R430AhAIF∆1−102ox specie, as WT, should undergo conformational

changes in order to get the dimer conformation.

52


BOOK OF ABSTRACTS

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Identification and development of new pharmacological

chaperones for phenylketonuria

M. Conde-Gimenez1,2, S. Salillas1,2, M. D. Dıaz-de-Villegas3 and J.

Sancho1,2.


Universidad de Zaragoza (Spain)2 BIFI and BIFI-IQFR(CSIC)-Joint Unit, Universidad de Zaragoza (Spain)3 Departamento de Catalisis y Procesos Catalıticos

Instituto de Sıntesis Quımica y Catalisis Homogenea (ISQCH)

Universidad de Zaragoza-CSIC (Spain)

Phenylketonuria (PKU) is an inherited metabolic disorder caused by mutations of pheny-

lalanine hydroxylase (PAH) gene. Nowadays, more than 550 disease-causing PAH mutations

have been identified and most of them induce loss of conformational stability and decreased

physiological enzymatic activity.

Currently, one of the most promising molecular approaches to treat this disease involves

using small molecules known as pharmacological chaperones that bind to the native state of

the unstable enzyme rescuing its physiological function. Actually, BH4, the natural cofactor of

PAH, is administered for the management of some PKU patients and it is believed that it acts

through that mechanism [1]. Despite this fact, BH4 is only effective for some forms of mild

PKU so that we propose to find novel PKU chaperones unrelated to BH4. A previous work

of this group identified two such compounds that enhanced the thermal stability of the WT

enzyme [2] and solved the X-ray structure of the complex between the enzyme and one of the

chaperones [3]. Based on these findings, we are synthetizing a group of novel compounds and

testing their effect on thermal denaturation assays. Concurrently, we are testing over 10,000

chemical compounds of a commercial diversity chemical library by applying a high-throughput

screening method. These new compounds may represent promising pharmacological entities

for non BH4 responsive PKU patients.

[1] Heintz C. et al. Hum Mut 2013, 34(7):927-36

[2] Pey A.L. et al. J Clin Invest 2008, 118(8): 2858-2867

[3] Torreblanca R. et al. ChemBioChem 2012, 13(9):1266-1269

53


BOOK OF ABSTRACTS

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Biochemical properties of the potential antimicrobial target

Ferric Uptake Regulator from Clostridium difficile

A. Fernandez-Otal1, M. L. Peleato2,3, M. Fillat2,3, A. Lanas1 and T.

Bes2,3

1 IIS Aragon-Instituto Aragones de ciencias de la Salud, Spain2 Departamento de Bioquımica y Biologıa Molecular y Celular,


Clostridium difficile is a strictly anaerobic Gram-positive bacillus that can be part of the

normal intestinal microbiota in healthy individuals. However, C. difficile is the main respon-

sible for antibiotic-associated colitis. Nowadays, this pathogen shows a considerable increase

in morbidity and mortality associated to severe C. difficile infections. In combination with

other strategies, the identification of bioactive compounds that may specifically inactivate

essential molecules of this microorganism and thereby inhibit or eradicate the infection could

lead to a rational design of new antimicrobials.

Fur (Ferric Uptake Regulator) belongs to a superfamily of regulatory proteins that control

iron homeostasis in many prokaryotes. This protein usually operates as a classical tran-

scriptional repressor preventing the transcription of Fur-regulated genes. Furthermore, this

DNA-binding protein regulates the expression of virulence genes in many pathogens under

iron-deficient conditions, a situation that occurs during the process of infection. Therefore,

Fur constitutes an attractive target for the development of new antimicrobial compounds.

As a first step to determine the suitability of C. difficile Fur protein (CdFur) as antibacte-

rial target, we have overexpressed in Escherichia coli as His-tagged protein and purified by

Immobilized Metal ion Affinity Chromatography the CdFur repressor. Gel retardation assays

(EMSA) have been used to verify the biological activity of the recombinant protein with se-

lected promoter genes under different conditions in vitro. Furthermore, a modified FURTA

(Fur titration assay) analysis has been set up in order to monitorize Fur activity in vivo.

54


BOOK OF ABSTRACTS

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Expresion genica de enzimas ligadas a la sınstesis de

melatonina en el tracto reproductor de morueco

A.E. Gaspar-Torrubia, D. Aguilar-Recarte, M Gonzalez-Arto, R

Perez-Pe, T Muino-Blanco, J.A.Cebrian-Perez and A Casao-Gascon

Department of Biochemistry and Molecular and Cellular Biology,

IUCA, Faculty of Veterinary, University of Zaragoza, Spain

La hormona melatonina es una de las sustancias biologicamente activas mas antiguas de los

organismos vivos, y su funcion principal es la defensa antioxidante. Su sıntesis es principal-

mente llevada a cabo por la glandula pineal durante la noche, y su concentracion en plasma

sanguıneo es practicamente nula durante el dıa.

Estudios llevados a cabo por nuestro grupo de investigacion han determinado su presencia

en plasma seminal ovino de manera constante y en, algunos momentos, en concentraciones

superiores a las detectadas en sangre. Este hecho podıa explicarse por la sıntesis de esta

hormona en el tracto reproductor del morueco, independiente de los ciclos de luz y oscuridad.

Con el fin de dilucidar la existencia de esta posible sıntesis en el tracto genital masculino, se

llevaron a cabo una serie de estudios en los distintos tejidos del aparato reproductor (testıculo,

epidıdimo y glandulas accesorias), centrados en demostrar la existencia de las dos enzimas

principales de la sıntesis: serotonina -N-acetiltransferasa (AANAT) y N-acetilserotonin-O-

metiltransferasa (ASMT). Mediante RT-PCR se demostro la expresion genica de dichas en-

zimas en todos los tejidos estudiados, y la q-PCR permitio comprobar que es mucho mayor

en el testıculo que en los demas tejidos. Por otra parte, mediante Western-blot se demostro

la traduccion y presencia de estas dos enzimas en todos los tejidos, lo que implica la exis-

tencia activa de un proceso de sıntesis de melatonina en el tracto reproductor del morueco,

principalmente en testıculo.

La importancia de dichos resultados radica en la existencia de la ruta de sıntesis de melaton-

ina y que su presencia en plasma seminal es un factor importante a la hora de determinar la

supervivencia y maduracion de las celulas espermaticas, aspecto fundamental en la capacidad

fecundante, maxime si se considera que esta hormona va a ser transferida al tracto genital

femenino tras la eyaculacion.

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BOOK OF ABSTRACTS

.

FurA influences heterocyst differentiation in Anabaena sp.

PCC 7120

A. Gonzalez1,2, A. Valladares3, T. Bes 1,2, M. L. Peleato1,2 and M.

Fillat1,2


Universidad de Zaragoza2 BIFI, Universidad de Zaragoza, Spain3 IBVF-CSIC-Universidad de Sevilla, Spain

In the filamentous nitrogen-fixing cyanobacterium Anabaena sp. PCC 7120, FurA is a global

transcriptional regulator whose expression is strongly induced by NtcA in proheterocysts

and remains stably expressed in mature heterocysts. In the present study, overexpression of

furA partially suppressed heterocyst differentiation by impairing morphogenesis at an early

stage. Recombinant purified FurA specifically bound in vitro to the promoter regions of ntcA,

while quantitative RT-PCR analyses indicated that furA overexpression strongly affected the

transient increase of ntcA expression that occurs shortly after nitrogen step-down. Overall,

the results suggest a connection between iron homeostasis and heterocyst differentiation via

FurA, by modulating the expression of ntcA.

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BOOK OF ABSTRACTS

.

Effect of mtDNA functional mutations on embryonic stem

cells gene expression

P. Meade1,2, S. Callejas3, N. Movilla1, P. Fernandez-Silva1,2 and J. A.

Enrıquez3.


Universidad de Zaragoza (Spain)2 BIFI and BIFI-IQFR(CSIC)-Joint Unit, Universidad de Zaragoza (Spain)3 Centro Nacional de Investigaciones Cardiovasculares CNIC. Madrid, Spain

Defects in the mitochondrial oxidative phosphorylation (OXPHOS) system lead to often

fatal, multi-system disorders affecting organs and tissues with a high-energy demand. Despite

the increasing knowledge in the underlying gene defects causing OXPHOS disorders, as well

as in their detection and diagnosis, the molecular pathogenesis events that link the mutated

gene to the observed clinical phenotypes are still largely unknown.

With the idea that mouse embryonic stem cells (ES) represent a more stable model than

the classical immortalized cell culture systems, we have used ES cells to asses the global

expression changes induced by alterations in the OXPHOS system. We have studied so far

three mtDNA mutations, generated in our lab, affecting the tRNAIle, ATPase6 and COX1

genes and causing different degrees of respiratory complex deficiency.

For this study, mitochondria harboring the mtDNA mutations were transferred from cul-

tured fibroblasts to C57BL6 ES cells, previously treated with rhodamine 6G. The transmi-

tochondrial ES cells clones were cultured for 9 passages and total RNA was extracted. The

global gene expression analysis was performed using Agilent mouse expression arrays.

Gene expression analysis revealed that in the transmitochondrial ES cells: a) rhodamine

6G treatment does not affect the pluripotency state of the ES cells; b) the changes observed

were mainly in genes participating in signalling pathways, as well as in inflammation and cell

communication processes; c) no significant changes were observed in carbohydrate and lipid

metabolism pathways.

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BOOK OF ABSTRACTS

.

TXNDC5, a newly discovered disulfide isomerase with a key

role in cell physiology and pathology

A. Pradilla Dieste1, C. Sanchez de Diego 1, E. Horna Terron1, J.

Osada2,3

1 Grado de Biotecnologıa. Universidad de Zaragoza2 Departamento de Bioquımica y Biologıa Molecular y Celular,

Facultad de Veterinaria, Universidad de Zaragoza3 Instituto deInvestigacion Sanitaria de Aragon (IIS), Universidad de Zaragoza

Thioredoxin domain containing 5 (TXNDC5) is a member of the protein disulfide isomerase

family. In fact, it boasts with Trx-like domains that catalyzes its thioredoxin activity and

allows it to act as a chaperone in the endoplasmic reticulum (ER) [1] Moreover, it can also work

as an electron transfer, recovering the functional isoform of other protein disulfide isomerases,

replacing reduced glutathione role [2,3]. Finally, it also acts as a cellular adapter, interacting

with N-terminal domain of adiponectin receptor [4,5]. As it can be inferred from all these

functions, TXNDC5 plays an important role in cell physiology; therefore, dysregulation of its

expression is associated with oxidative stress, cell ageing and a large range of pathologies such

as diabetes [6], arthritis [7-9], cancer [10], neurodegenerative diseases [11], and vitiligo [12]. For

instance, it has been found that TXNDC5 is upregulated in hepatocellular carcinoma [13,14],

esophagus, liver, cervix [15], uterus, colon, stomach [16-18], prostate, pancreas [17,19], renal

tumor tissue [20] and myeloma cells. However, hypoxia does not always induce TXNDC5

up regulation. For example, in non-small cell lung carcinoma, hypoxia does not change

TXNDC5 expression [17,19] [16] [20]. Furthermore, the presence of the Conjoined Gene

(CG) MUTED-TXNDC5 reduces TXNDC5 expression without modifying TXNDC5 mRNA

levels in a significant way [21]. These findings indicate that the expression and function of

TXNDC5 may be complex and variable according to tissue and cell conditions. However, its

role in the cell, as well as, its implication in all those important diseases has turned TXNDC5

into a susceptible biomarker or even a potential pharmacological target.

[1] Crown Human Genome Center DoMG, the Weizmann Institute of Science Thioredoxin

domain containing 5 2014,

[2] Araki K, Iemura S, Kamiya Y, Ron D, Kato K, Natsume T, Nagata K: Ero1-alpha and

pdis constitute a hierarchical electron transfer network of endoplasmic reticulum oxidoreduc-

tases. J Cell Biol 2013;202:861-874.

[3] Shepherd C, Oka OB, Bulleid NJ: Inactivation of mammalian ero1alpha is catalysed by

specific protein disulfide-isomerases. Biochem J 2014;461:107-113.

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[4] Charlton HK, Webster J, Kruger S, Simpson F, Richards AA, Whitehead JP: Erp46

binds to adipor1, but not adipor2, and modulates adiponectin signalling. Biochem Biophys

Res Commun 2010;392:234-239.

[5] Heiker JT, Kosel D, Beck-Sickinger AG: Molecular mechanisms of signal transduction

via adiponectin and adiponectin receptors. Biol Chem 2010;391:1005-1018.

[6] Chen DL, Xiang JN, Yang LY: Role of erp46 in beta-cell lipoapoptosis through endoplas-

mic reticulum stress pathway as well as the protective effect of exendin-4. Biochem Biophys

Res Commun 2012;426:324-329.

[7] Chang X, Cui Y, Zong M, Zhao Y, Yan X, Chen Y, Han J: Identification of proteins with

increased expression in rheumatoid arthritis synovial tissues. J Rheumatol 2009;36:872-880.

[8] Chang X, Zhao Y, Yan X, Pan J, Fang K, Wang L: Investigating a pathogenic role for

txndc5 in rheumatoid arthritis. Arthritis Res Ther 2011;13:R124.

[9] Wang L, Zheng Y, Xu H, Yan X, Chang X: Investigate pathogenic mechanism of txndc5

in rheumatoid arthritis. PLoS One 2013;8:e53301.

[10] Camargo LL, Babelova A, Mieth A, Weigert A, Mooz J, Rajalingam K, Heide H, Wittig

I, Lopes LR, Brandes RP: Endo-pdi is required for tnfalpha-induced angiogenesis. Free Radic

Biol Med 2013;65:1398-1407.

[11] Lin SH, Liu CM, Liu YL, Shen-Jang Fann C, Hsiao PC, Wu JY, Hung SI, Chen CH,

Wu HM, Jou YS, Liu SK, Hwang TJ, Hsieh MH, Chang CC, Yang WC, Lin JJ, Chou FH,

Faraone SV, Tsuang MT, Hwu HG, Chen WJ: Clustering by neurocognition for fine mapping

of the schizophrenia susceptibility loci on chromosome 6p. Genes Brain Behav 2009;8:785-794.

[12] Jeong KH, Shin MK, Uhm YK, Kim HJ, Chung JH, Lee MH: Association of txndc5

gene polymorphisms and susceptibility to nonsegmental vitiligo in the korean population. Br

J Dermatol 2010;162:759-764.

[13] Park MS, Kim SK, Shin HP, Lee SM, Chung JH: Txndc5 gene polymorphism contributes

to increased risk of hepatocellular carcinoma in the korean male population. Anticancer Res

2013;33:3983-3987.

[14] Nissom PM, Lo SL, Lo JC, Ong PF, Lim JW, Ou K, Liang RC, Seow TK, Chung MC:

Hcc-2, a novel mammalian er thioredoxin that is differentially expressed in hepatocellular

carcinoma. FEBS Lett 2006;580:2216-2226.

[15] Chang X, Xu B, Wang L, Wang Y, Yan S: Investigating a pathogenic role for txndc5

in tumors. Int J Oncol 2013;43:1871-1884.

[16] Wang Y, Ma Y, Lu B, Xu E, Huang Q, Lai M: Differential expression of mimecan

and thioredoxin domain-containing protein 5 in colorectal adenoma and cancer: A proteomic

study. Exp Biol Med (Maywood) 2007;232:1152-1159.

[17] Zhang L, Hou Y, Li N, Wu K, Zhai J: The influence of txndc5 gene on gastric cancer

cell. J Cancer Res Clin Oncol 2010;136:1497-1505.

[18] Zhang L, Hou YH, Wu K, Zhai JS, Lin N: Proteomic analysis reveals molecular biologi-

cal details in varioliform gastritis without helicobacter pylori infection. World J Gastroenterol

2010;16:3664-3673.

[19] Vincent EE, Elder DJ, Phillips L, Heesom KJ, Pawade J, Luckett M, Sohail M, May

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MT, Hetzel MR, Tavare JM: Overexpression of the txndc5 protein in non-small cell lung

carcinoma. Anticancer Res 2011;31:1577-1582.

[20] Duivenvoorden WC, Paschos A, Hopmans SN, Austin RC, Pinthus JH: Endoplasmic

reticulum protein erp46 in renal cell carcinoma. PLoS One 2014;9:e90389.

[21] Bioinformatics LfI: Conjoing: Database of conjoined genes; in RIKEN (ed, 2014, 2014,

pp Database for Conjoined Genes.

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BOOK OF ABSTRACTS

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Development of compounds with promising selective effects

toward Helicobacter pylori flavodoxin

S. Salillas1,2, M. Alias2, A. Velazquez-Campoy1,2,3, J. A. Carrodeguas1,2,

M. D. Dıaz-de-Villegas4 and J. Sancho1,2.


Universidad de Zaragoza (Spain)2 BIFI and BIFI-IQFR(CSIC)-Joint Unit, Universidad de Zaragoza (Spain)3 Fundacion ARAID, Gobierno de Aragon. Zaragoza (Spain)4 Departamento de Catalisis y Procesos Catalıticos

Instituto de Sıntesis Quımica y Catalisis Homogenea (ISQCH)

Universidad de Zaragoza-CSIC (Spain)

Helicobacter pylori (Hp) establishes life-long infections in the gastric mucosa of humans that

may develop in a diversity of diseases from ulcer to cancer. Rising resistance of the bacteria

to the antibiotics used in current treatments makes necessary the study of new therapies

[1]. A specific target of this bacteria is its flavodoxin protein (Hp-Fld), that is essential for

its life. This protein exhibits a distinct pocket near the cofactor binding site, which could

interact with selective organic compounds interfering with flavodoxin function2. We have

recently developed, through organic synthesis, novel potential therapeutics based on one of

the three molecules that we identified previously as flavodoxin inhibitors [2]. We expected

these compounds to bind flavodoxin with a certain association constant, that is why we have

monitored the binding of the inhibitors to Hp-Fld by isothermal titration calorimetry (ITC).

In addition, we have tested these four novels analogous for cytotoxicity toward HeLa cells

using both microscopy of living cells and cytotoxicity assays (XTT method). All of the new

molecules show less toxicity toward HeLa cells than the original lead compounds. To establish

their bactericidal effect, efficacy assays are currently in progress. These trials will allow us to

establish whether these new compounds could represent promising selective inhibitors of the

Helicobacter pylori flavodoxin, with the additional advantage of not generally perturbing the

bacterial flora and not being yet counteracted by Hp resistance mechanisms.

[1] Cremades N et al. ACS Chem. Biol. 2009, 4: 928-938.

[2] Galano J et al. J. Med. Chem. 2013, 56: 6248-6258.

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Key residues regulating the reductase activity of the human

mitochondrial Apoptosis Inducing Factor

R. Villanueva1,2, C. Marcuello3, A. Uson1,2, M. D. Miramar1, M. L.

Peleato1,2, A Lostao3,4, S. Susin5, M. Medina1,2 and P. Ferreira1,2


Universidad de Zaragoza2 BIFI, Universidad de Zaragoza, Spain3 Laboratorio de Microscopias Avanzadas, INA, Universidad de Zaragoza, Spain4 Fundacion ARAID, Zaragoza, Spain5 INSERM U1138, Centre de Recherche des Cordeliers,

Unviersite de Paris Descartes-Sorbonne, Paris, France

The human Apoptosis Inducing Factor (hAIF) is a bifunctional NAD(P)H-dependent fla-

voreductase involved in mitochondrial energy metabolism and caspase-independent cell death.

Many studies indicate that both functions are redox controlled by NADH binding, although

the exact role of hAIF as a reductase in healthy mitochondria remains still unknown. Upon

reduction by NADH, hAIF dimerizes and forms very stable FAD:nicotinamide charge transfer

complexes (CTC) that contribute to restrict its efficiency as a reductase. The molecular bases

of the hAIF reductase activity were investigated by analyzing the role played by particular

residues at the isoalloxazine environment of its FAD cofactor. We observed that replacements

of K177 and E314 have drastic effects in hAIF ability to retain the FAD cofactor as well as

to maintain the reduced protein in complex with the reaction product (NAD+). The stacking

interaction established by P173 with the isoalloxazine ring seems also critical to determine

the flavin environment and to modulate the enzyme affinity for NADH. Characterization of

mutations located at residues F310 and H454, indicates that these two positions contribute

to form a compact active site essential for NADH binding, CTC stabilization, and NAD+

affinity for the reduced state of the protein. Altogether, we demonstrate that these catalytic

features are the key to determine the particular behavior of hAIF as a reductase.

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