protein functions prediction

Swiss Institute of BioinformaticsInstitut Suisse de Bioinformatique

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Protein functions prediction


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Introduction

Signal peptides Transmembrane

regions and topology PTM (post-

translational modifications)

Low complexity and biased regions

Repeats Coils

Secondary structure Antigenic peptides Domain/Motifs Tools The EMBOSS package


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Different techniques

Algorithms Sliding window, Nearest Neighbor Patterns, regular expression Weight matrices HMM, profiles Neural Networks Rules


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Sliding window

THISISATESTSEQVENCETHATDISPLAYSTHESLIDINGWINDQW

Score1Score2

Scoren

Width or Size=11, Step=5

Results are usually displayed as a graph, see example ->


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Patterns / regular expression

Pattern: <A-x-[ST](2)-x(0,1)-{V} Regexp: ^A.[ST]{2}.?[^V] Text: The sequence must start with an

alanine, followed by any amino acid, followed by a serine or a threonine, two times, followed by any amino acid or nothing, followed by any amino acid except a valine.

Simply the syntax differ…


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Weight matrices (PSSM)


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HMM / profiles


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Neural Networks

General principle: Example:


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Signals found in proteins

N-ter exportation - secretion mitochondria chloroplast

internal NLS (nuclear

localization signal)

C-ter GPI-anchor (Glycosyl

Phosphatidyl Inositol) other membrane

anchors (see PTM) other unknown ?


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Signals detection tools

SignalP MitoProt ChloroP Predotar PSort TargetP Sigcleave (EMBOSS) Phobius

Big-PI DGPI


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Transmembrane regions

Detection (signal peptide, hydropathy, helices) Organisation (topology)


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Transmembrane detection tools

TMHMM TMPred TopPred2 DAS HMMTop Tmap (EMBOSS)

Mixture of tools Phobius ConPred II


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Post translational modifications

Phosphorylation S - T - Y

N-glycosylation N

O-glycosylation S - T - (HO)K

Acetylation, methylation D - E - K

Sulfation Y

Farnesylation, myristylation, palmitoylation, geranylgeranylation, GPI-anchor C - Nter - Cter

Ubiquitination and family K - Nter

Inteins (protein splicing) Pre-translational

Selenoprotein C


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PTM detection

Pattern prediction (PROSITE)

Short or weak signal Frequent hit producer Best method is

experimental MS/MS detection

Most method use « rules » joining pattern detection and knowledge to predict sites.

NetOGlyc - Prediction of type O-glycosylation sites in mammalian proteins

DictyOGlyc - Prediction of GlcNAc O-glycosylation sites in Dictyostelium

YinOYang - O-beta-GlcNAc attachment sites in eukaryotic protein sequences

NetPhos - Prediction of Ser, Thr and Tyr phosphorylation sites in eukaryotic proteins

NMT - Prediction of N-terminal N-myristoylation

Sulfinator - Prediction of tyrosine sulfation sites


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Low complexity regions

repeats compositional bias PEST


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Low complexity / Repeats

DUST (DNA) / SEG de novo detection

RepeatMasker (DNA) search collection

REP search collection

REPRO, Radar de novo detection

PEST, PESTFind de novo detection

EMBOSS (DNA) einverted equicktandem etandem palindrome

EMBOSS (protein) oddcomp


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Coils

Helix of helix coiled-coil

Leu-zipper


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Coils detection

COILS Weight matrices

Paircoil, Multicoil Pairwise correlation

Marcoil HMM

Pepcoil (EMBOSS) Weight matrices


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Secondary structure

Structure to predict Alpha-helices Beta-sheets Turns Random coil

Garnier (EMBOSS) PHD DSC PREDATOR NNSSP Jpred Jnet Many others


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Antigenic peptide

Peptides binding to MHC class I

8, 9, 10 mers class II

15 mers (3+9+3) Depend highly on MHC

type

Use of experimental knowledge Databases of known

peptides

SYFPEITHI HLA_Bind (BIMAS) MAPPP combined expert Antigenic (EMBOSS) Many more

Prediction of proteasome cleavage sites

NetChop PaProc


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Domain / Motif

All the protein domain descriptors PROSITE PFAM SMART PRODOM BLOCKS PRINTS TIGRfam …

Federation: InterPro Many techniques

Patterns, Regexp PSSM (PSI-BLAST) Profiles HMM


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Other Tools

You can find some of them on our servers www.ch.embnet.org

Or on ExPASy server www.expasy.org/tools

Or ask Google!! www.google.com


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European Molecular Biology Open Software Suite


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How to use EMBOSS/Jemboss at SIB


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Free Open Source (for most Unix plateforms) GCG successor (compatible with GCG file format) More than 150 programs (ver. 2.9.0) Easy to install locally

but no interface, requires local databases Unix command-line only

Interfaces Jemboss, www2gcg, w2h, wemboss… (with account) Pise, EMBOSS-GUI, SRSWWW (no account) Staden, Kaptain, CoLiMate, Jemboss (local)

Access: www.emboss.org or emboss.sourceforge.net


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Format USA 'asis' :: Sequence [start : end : reverse] Format :: '@' ListFile [start : end : reverse] Format :: 'list' : ListFile [start : end : reverse] Format :: Database : Entry [start : end : reverse] Format :: Database - SearchField : Word [start : end : reverse] Format :: File : Entry [start : end : reverse] Format :: File : SearchField : Word [start : end : reverse] Format :: Program Program-parameters '|' [start : end : reverse]

Example: fasta::Swissprot:UBP5_HUMAN[200:300]

Databases Any can be added, use showdb to display the available databases

Some details


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showdbDisplays information on the currently available databases# Name Type ID Qry All Comment# ==== ==== == === === =======ipr_fetch P OK OK OK InterPro current by fetchipi_fetch P OK OK OK IPI current by fetchrefseq_fetch P OK OK OK refseq current by fetchrepbase_fetch P OK OK OK repbase current by fetchswiss_fetch P OK OK OK SwissProt current by fetchswissprot P OK OK OK SWISSPROT sequencestrembl P OK OK OK TREMBL sequencestrembl_fetch P OK OK OK trembl current by fetchtremblnew P OK OK OK TREMBL New sequencesug_fetch P OK OK OK Unigene by fetchembl N OK OK OK EMBL releaseemhum N OK OK OK EMBL release, Human section by emboss indexemrod N OK OK OK EMBL release, Rodent section by emboss indexemvrt N OK OK OK EMBL release, Vertebrate (nonhuman, nonrodent)

seqret (seqretall, seqretset, seqretsplit) entret (for complete untouched entry, e.g., for unigene, interpro,

swissprot…) Possible to define your own « .embossrc » file

databases


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Some tools for DNA redata Search REBASE for enzyme name, references, suppliers etc remap Display a sequence with restriction cut sites, translation etc restover Finds restriction enzymes that produce a specific overhang restrict Finds restriction enzyme cleavage sites showseq Display a sequence with features, translation etc silent Silent mutation restriction enzyme scan cirdna Draws circular maps of DNA constructs lindna Draws linear maps of DNA constructs revseq Reverse and complement a sequence …


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Example: remap

ECLAC E.coli lactose operon with lacI, lacZ, lacY and lacA genes. Hin6I TaqI | HhaI | Bsc4I | Bsu6I | | Hin6I | BssKI | | | HhaI AciI | | BsiSI \ \ \ \ \ \ \ \ GACACCATCGAATGGCGCAAAACCTTTCGCGGTATGGCATGATAGCGCCCGGAAGAGAGT 10 20 30 40 50 60 ----:----|----:----|----:----|----:----|----:----|----:----| CTGTGGTAGCTTACCGCGTTTTGGAAAGCGCCATACCGTACTATCGCGGGCCTTCTCTCA / / / / / / / /// | TaqI | Hin6I AciI | | ||BssKI Bsc4I HhaI | | |BsiSI | | Bsu6I | Hin6I HhaI# Enzymes that cut Frequency Isoschizomers AciI 1 Bsc4I 1 BsiSI 1 BssKI 1 Bsu6I 1 HhaI 2 Hin6I 2 HinP1I,HspAI TaqI 1# Enzymes that do not cutAclI BamHI BceAI Bse1I BshI ClaI EcoRI EcoRII Hin4I HindII HindIII HpyCH4IV KpnI NotI


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Example: cirdna File: ../../data/data.cirpStart 1001End 4270grouplabelBlock 1011 1362 3ex1endlabellabelTick 1610 8EcoR1endlabellabelBlock 1647 1815 1endlabellabelTick 2459 8BamH1endlabellabelBlock 4139 4258 3ex2endlabelendgroupgrouplabelRange 2541 2812 [ ] 5AluendlabellabelRange 3322 3497 > < 5MER13endlabelendgroup


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Example: plotorf


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EMBOSS format input/output

UFO Universal Feature Object gff, swissprot, embl, pir, nbrf (with or without sequence)

Alignments Multiple and pairwise, many flavors (FASTA, MSF, SRS…)

Reports Feature (UFO), SRS, motif, seqtable, excel, diffseq, listfile

(USA), etc… Sequences (compatible with USA)

Many!!! E.g., fasta, clustal, gcg, paup, gff, embl, swissprot, acedb, abi, etc…


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Web interfaces

PISE (Pasteur Institute Software Environment) http://www-alt.pasteur.fr/~letondal/Pise/

wEMBOSS (Belgium&Argentina) (not yet at SIB) http://www.wemboss.org


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Pise a tool to generate Web interfaces for Molecular Biology programs

http://emboss.ch.embnet.org/Pise


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http://www.wemboss.org


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Launch Jembosshttp://emboss.ch.embnet.org/Jemboss


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Launch Jemboss

First time only…

Each time…


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Jemboss windows


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Jemboss windows other systems


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Summary

Anonymous web access through Pise Registered access through Jemboss Registered access through command-line

(requires UNIX skills)

Please report problems!


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Exercises

DEA Exercises web based sequence analysis The goal of this exercise is to use web based tools for protein sequence analysis

a) Take this TrEMBL sequence (Q9X252) and try a BLAST against swissprot with the complete protein or with the first 70 residues. Explain the difference. Use TMPred, SignalP, and COILS to help you.

b) Pass this sequence through PFSCAN and search all databases. Compare with this command on ludwig-sun1/2: hits -b "prf pat pfam" tr:Q9X252

c) use the different profile, motifs, pattern databases to get more information about the domain(s) you found.

d) How do you evaluate the PRINTS tropomyosin annotation in this TrEMBL entry (Q9WZH0)? List of useful links:

basic BLAST or advanced BLAST or PSI-BLAST TMPred prediction tool for transmembrane regions (or TMHMM) COILS prediction tool for coiled-coil regions SignalP prediction tool for signal-peptide cleavage site

Profile, domain, motifs databases and search sites: PFSCAN InterPro (Pfam, PRINTS, PROSITE, SMART) HITS

protein functions prediction

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