Basic bioinformatics tools for studying proteins

Basic bioinformatics tools for studying proteins

Dong Xu

Computer Science Department C. S. Bond Life Sciences CenterUniversity of Missouri, Columbia

http://digbio.missouri.edu

Introduction

Broaden knowledge for undergraduate education

Many opportunities for biomedical and agricultural related jobs

Practice basic protein tools:Useful for biological studiesIntellectually stimulating

Dong’s picks for beginners :Not unnecessarily the most accurate toolEasy to use and understandVery popular

Proteins – Some Basics

What Is a Protein?Linear Sequence of Amino Acids...

What is an Amino Acid?

20 20 Amino acidsAmino acids

Glycine (G)

Glutamic acid (E)

Asparatic acid (D)

Methionine (M)

Threonine (T)

Serine (S)

Glutamine (Q)

Asparagine (N)

Tryptophan (W)

Phenylalanine (F)

Cysteine (C)

Proline (P)

Leucine (L)

Isoleucine (I)

Valine (V)

Alanine (A)

Histidine (H)

Lysine (K)

Tyrosine (Y)

Arginine (R)

White: Hydrophobic, Green: Hydrophilic, Red: Acidic, Blue: Basic

Amino Acids connect via PEPTIDE BOND

Peptide Bond

A AFNG

GS T

SD

K

An Overview

o A protein folds into a unique 3D structure under the physiological condition

Lysozyme sequence (129 amino acids):KVFGRCELAA AMKRHGLDNY RGYSLGNWVC AAKFESNFNT QATNRNTDGS

TDYGILQINS RWWCNDGRTP GSRNLCNIPC SALLSSDITA SVNCAKKIVS

DGNGMNAWVA WRNRCKGTDV QAWIRGCRL

Protein backbones: Side chain

Primary, Secondary and Tertiary Structures of

Proteins

Protein Structure Representations

Lysozyme structure:

ball & stick strand surface

Structure Visualization

Rasmol (http://www.umass.edu/microbio/rasmol/getras.htm)

MDL Chime (plug-in) (http://www.mdl.com/products/framework/chime/)

Protein Explorer (http://molvis.sdsc.edu/protexpl/frntdoor.htm)

Jmol: http://jmol.sourceforge.net/ Pymol: http://pymol.sourceforge.net/ Vmd: http://www.ks.uiuc.edu/Research/vmd/

Sequence Homology Software

NCBI-BLASThttp://www.ncbi.nlm.nih.gov/BLAST/

Comparing 2 (pairwise) or more (multiple) sequences.

Searching for a series of identical or similar characters in the sequences.

VLSPADKTNVKAAWAKVGAHAAGHG||| | | |||| | ||||VLSEAEWQLVLHVWAKVEADVAGHG

Typical BLAST Output

InterPro Scanhttp://www.ebi.ac.uk/InterProScan/

InterPro Scan PCNA http://www.ebi.ac.uk/InterProScan/

MyHits Local Motifs Searchhttp://myhits.isb-sib.ch/

MyHits Local Motifs Summaryhttp://myhits.isb-sib.ch/

MyHits Local Motif Hitshttp://myhits.isb-sib.ch/

Multiple Alignment

VTISCTGSESNIGAG-NHVKWYQQLPGVTISCTGTESNIGS--ITVNWYQQLPGLRLSCSSSDFIFSS--YAMYWVRQAPGLSLTCTVSETSFDD--YYSTWVRQPPGPEVTCVVVDVSHEDPQVKFNWYVDG--ATLVCLISDFYPGA--VTVAWKADS--AALGCLVKDYFPEP--VTVSWNSG---VSLTCLVKEFYPSD--IAVEWWSNG--

Phylogeny Tree

Multiple protein sequence alignment

conserved sites and hence possibly functional sites

phylogenetic tree

MSA with ClustalW

1exr_A -EQLTEEQIAEFKEAFALFDKDGDGTITTKELGTVMRSLGQNPTEAELQDMINEVDADGN 59 1N0Y_A AEQLTEEQIAEFKEAFALFDKDGDGTITTKELGTVMRSLGQNPTEAELQDMINEVDADGN 60 3cln_ ----TEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRSLGQNPTEAELQDMINEVDADGN 56 :************:******************************************* 1exr_A GTIDFPEFLSLMARKMKEQDSEEELIEAFKVFDRDGNGLISAAELRHVMTNLGEKLTDDE 119 1N0Y_A GTIDFPEFLSLMARKMKEQDSEEELIEAFKVFDRDGNGLISAAELRHVMTNLGEKLTDDE 120 3cln_ GTIDFPEFLTMMARKMKDTDSEEEIREAFRVFDKDGNGYISAAELRHVMTNLGEKLTDEE 116 *********::******: *****: ***:***:**** *******************:* 1exr_A VDEMIREADIDGDGHINYEEFVRMMVS- 146 1N0Y_A VDEMIREADIDGDGHINYEEFVRMMVSK 148 3cln_ VDEMIREANIDGDGQVNYEEFVQMMTA- 143 ********:*****::******:**.:

2 or more sequences for

analysis

params (default or custom for

different scoring

matrices, gap penalties, etc.)

ClustalW

Phylogram

Cladogram

ClustalW: http://www.ebi.ac.uk/Tools/clustalw2/index.html

Cell localization

Typical Sorting Signals

Signal Function Example

Import into nucleus -P-P-K-K-K-R-K-V-

Export from nucleus -L-A-L-K-L-A-G-L-D-I-

Import into mitochondria <-MLSLRQSIRFFKPATRTLCSSRYLL-

Import into plastid <-MVAMAMASLQSSMSSLSLSSNS

FLGQPLSPITLSPFLQG-

Import into peroxisomes -S-K-L->

Import into ER <-MMSFVSLLLVGILFWAT

EAEQLTKCEVFN-

Return to ER -K-D-E-L->

Localizations

Cell localization

PSORT: http://psort.nibb.ac.jp/

TargetP:

http://www.cbs.dtu.dk/services/TargetP/

Signal peptide

SingalP:

http://www.cbs.dtu.dk/services/SignalP/

SignalP result

Membrane Bilayer with Proteins

Helix Bundle TM Proteins

PDB = 1QHJ PDB = 1RRC

Single helix or helical bundles (> 90% of TM proteins)Examples: Human growth hormone receptor, Insulin receptor

ATP binding cassette family - CFTRMultidrug resistance proteins

7TM receptors - G protein-linked receptors

Beta Barrel TM Proteins

Transmembrane Prediction

http://bp.nuap.nagoya-u.ac.jp/sosui/ (alpha)

http://psfs.cbrc.jp/tmbeta-net/ (beta)

Secondary Structure Prediction

SSpro 4.1: http://sysbio.rnet.missouri.edu/multicom_toolbox/

PSI-PRED: http://bioinf.cs.ucl.ac.uk/psipred/psiform.html

SAM: http://compbio.soe.ucsc.edu/SAM_T08/T08-query.html

PHD: http://www.predictprotein.org/

Coiled coil prediction

http://npsa-pbil.ibcp.fr/cgi-bin/npsa_automat.pl?page=/NPSA/npsa_lupas.html

Special motif prediction

Helix-turn-helix motif predictionhttp://npsa-pbil.ibcp.fr/cgi-bin/npsa_automat.pl?page=/NPSA/npsa_hth.html

Kinase related motifshttp://scansite.mit.edu/motifscan_seq.phtml

Leucine Zippershttp://2zip.molgen.mpg.de/index.html

Protein disorder prediction

PreDisorder: http://sysbio.rnet.missouri.edu/multicom_toolbox/

A collection of disorder predictors:http://www.disprot.org/predictors.php

2D: Contact Map Prediction

1 2 ………..………..…j...…………………..…n 123....i.......n

3D Structure 2D Contact Map

Distance Threshold = 8Ao

Contact Prediction

SVMcon: http://casp.rnet.missouri.edu/svmcon.html NNcon:

http://casp.rnet.missouri.edu/nncon.html SCRATCH: http://scratch.proteomics.ics.uci.edu/ SAM:

http://compbio.soe.ucsc.edu/HMM-apps/HMM-applications.html

Structure Comparison

Visualize structure alignment using VAST:

http://www.ncbi.nlm.nih.gov/Structure/

Two ferredoxins, 1DOI and

1AWD, are aligned structurally,

showing an insertion in 1DOI

that contains potassium-ion

binding sites. This may be the

result of adaptations to the high

salt environment of the Dead Sea.

Structure Alignment Tools

CE (http://cl.sdsc.edu/) DALI

(http://www.ebi.ac.uk/dali/)

TM-Align: http://zhang.bioinformatics.ku.edu/TM-align/

Structure-Based Search

Comparing a query protein structure against

all the structures in the PDB

The DALI server:

http://www2.ebi.ac.uk/dali/

When new structures are solved, researchers often submit them to the DALI server to find structural neighbors and their alignments.

Swiss Model: Comparative Modeling Serverhttp://swissmodel.expasy.org/

Protein Structure Homology Modeling: Modeller

Analysis software

PROCHECK WHATCHECK Suite Biotech PROSA

Entrez Databaseshttp://www.ncbi.nlm.nih.gov/Entrez/

Design Program

DEZYMER (Hellinga)Given a ligand and a protein with known structure,

suggest residues to be mutated so that the resulting protein binds the ligand.

ORBIT (Mayo)Given a backbone structure, design a sequence such

that it folds to that backbone.

Rosetta (Baker)One program to treat diverse problems

Prediction and design

DEZYMER

1. Define the expected binding geometry

2. Find backbone places where if appropriate side chains are added, the predefined geometry is satisfied

3. Place the side chains and ligand, and optimize there position

4. Repack residues in positions other than binding residues. If necessary, change residue type

Hellinga and Richards, JMB, 1991. Construction of new ligand binding sites in protein of known structure

ORBIT

Comparison between the designed backbone (averaged NMR structure, blue) and the target backbone (red)

Solution structure of the designed protein. Stereoview showing the best-fit superposition of the 41

1. Divide the target structure into three parts: core, surface and boundary

2. Core: Ala, Val, Leu, Ile, Phe, Tyr, Trp Surface: Ala, Ser, Thr, His, Asp, Asn, Glu, Gln, Lys, and Arg Boundary: union of the above two

3. 1.9*1027 possible sequence

4. Select best sequence efficiently, using dead end elimination (DDE)

Calciomics

Calciomics is a specialized area of biochemistry focusing on the study of calcium-binding biological macromolecules and proteins to understand the factors that contribute to calcium-binding affinity and the selectivity of proteins and calcium-dependent conformational change.

http://lithium.gsu.edu/faculty/Yang/Calciomics.htm

SOSUIRemove transmembrane

regions

SignalPRemove signal region

ProDom

Modifiedsequences

PROSPECT

Originalsequence

Set of domainsequences

Coiled coilsRemove disorder

regions

SSPSecondary Structure

prediction

PSI-BLAST

Iterations:Analysis of E-value,

set of profile sequences

STOPif homolog

found in PDB

3D model

Function annotation

SWISS-PROTannotation

PFAMFamily classification

MotifActive sites

PSORTSubcellular location

Enzyme structure DB

MedlineLiterature search

WHATIF /PROCHECK

Evaluate & adjust alignments

MODELLER/ Jackle

seq

uen

ce

anal

ysis

and

pro

cess

ing

stru

ctu

re p

red

icti

on a

nd

eva

luat

ion

fun

ctio

n in

fere

nce

tool

kit

Summary

Practice 10 selected tools Help answer the question: what does

this protein do? Collaborate with experimentalists Find more tools at

http://us.expasy.org/tools/http://infosuite.welch.jhmi.edu/BS/pt

Acknowledgments

This file is for the educational purpose only. Some materials (including pictures and text) were taken from the Internet at the public domain.