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From Genes to Proteins(and a little bit about allignments)

Prof:Rui Alvesralves@cmb.udl.es

973702406Dept Ciencies Mediques Basiques,

1st Floor, Room 1.08Website of the

Course:http://web.udl.es/usuaris/pg193845/Courses/Bioinformatics_2007/ Course: http://10.100.14.36/Student_Server/

(and a little about allignments)

• How does blast work?

How Does BLAST Really Work?

• The BLAST programs improved the overall speed of searches while retaining good sensitivity (important as databases continue to grow) by breaking the query and database sequences into fragments ("words"), and initially seeking matches between fragments.

BLAST Algorithm

How Does BLAST Really Work?

• Word hits are then extended in either direction in an attempt to generate an alignment with a score exceeding the threshold of "S".

BLAST Algorithm

Extending the High Scoring Segment Pair (HSP)

NeighborhoodScore Threshold

MinimumScore

SignificanceDecay

BLAST Algorithm

• Sequences are split into words (default n=3)– Speed, computational efficiency

• Scoring of matches done using scoring matrices• HSP = high scoring segment pair

– BLAST algorithm extends the initial “seed” hit into an HSP

• Local optimal alignment• More than one HSP can be found

Where does the score (S) come from?

• The quality of each pair-wise alignment is represented as a score and the scores are ranked.

• Scoring matrices are used to calculate the score of the alignment base by base (DNA) or amino acid by amino acid (protein).

• The alignment score will be the sum of the scores for each position.

Organization of the Talk

• From cDNA sequence to protein sequence.

Predicting protein sequence from DNA sequence

• Protein sequence can be predicted by translating the cDNA and using the genetic code.

Translating cDNA into protein sequence

ATGTCTCTTATATGA…

MetSerLeuIleTer

No Gene!!!!!

Sometimes stop codons are not stop codons.

Are there alternative genetic codes?

• YES!!!!

Codon Universal code

AAA K N(9,14)

AAC K

AAG N

AAT N

ACA T

ACC T

ACG T

ACT T

AGA R END(2) S(5,9,14,21) G(13)

AGC S

AGG R END(2) S(5,9,14,21) G(13)

AGT S

ATA I M(2,3,5,13,21)

ATC I

ATG M

ATT I

Echinoderm/Flatworm mitochondria

Flatworm mitochondria

Vertebrate mitochondria

Yeast mitochondria Invertebrate

mitochondria

Ascidian mitochondria

Trematode mitochondria

Codon Universal code

CAA Q

CAC H

CAG Q

CAT H

CCA P

CCC P

CCG P

CCT P

CGA R

CGC R

CGG R

CGT R

CTA L T(3)

CTC L T(3)

CTG L T(3) S(12)

CTT L T(3)

Yeast mitochondria

Candida

Codon Universal code

GAA E

GAC D

GAG E

GAT D

GCA A

GCC A

GCG A

GCT A

GGA G

GGC G

GGG G

GGT G

GTA V

GTC V

GTG V

GTT V

Codon Universal code

TAA END Q(6) Y(14)

TAC Y

TAG END Q(6,15) L(16,21)

TAT Y

TCA S END(21)

TCC S

TCG S

TCT S

TGA END W(2,3,4,5,6,9,13,14,21) C(10)

TGC C

TGG W

TGT C

TTA L

TTC F

TTG L

TTT F

Ciliate, Dasycladacean Hexamita

Translating yeast mitochondrial cDNA into protein sequence

ATGTCTCTTATATGA………SECIS sequence

TrpSerThrMetsCys

MetSerLeuIleTer

There is a Gene with a considerably different protein sequence from the one we would

predict from the universal genetic code!!!!!

What changed?

• END→AA (16 events, all ter codons)

• AA →END (3 events)

• AA →AA’ (22 events, 3 AA)

Conclusions

• Translating DNA or mRNA sequence into proteins is easy but you need to be carefull as to which genetic code you need to use.