homo logy modeling

8/2/2019 Homo Logy Modeling

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OutlineOutline Introduction

What is homology modeling

Theory Basic principles and procedure

InsightII and homology module How to start it

Building your first model from sequence

Dihydrofolate Reductase Improving your model

Relaxation


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IntroductionIntroduction

Goal

Create a 3-D structure of a protein Why

When the 3-D structure is not available from

X-ray or NMR studies What do you need to know

Protein sequence of interest (target sequence)

Protein structure of at least one homologousprotein (3-D template)


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What is a homologous protein

A protein that have a high sequence similarityto the target protein sequence

Basic assumption of Homology Modeling Proteins sharing high sequence similarity

should have similar protein fold

. . . . (if they dont misfold)


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IntroductionIntroduction There are exceptions !!

Protein can be designed with a fewmutations/insertions/deletions to completelyunfold, misfold and even fold to another

secondary structure

Proteins may adopt completely different folds

depending on environmental variables such aspH, temperature, salt conc. etc. which is notaddress by homology modeling


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Why are we still doing it ??

If someone ask you, Given a class of proteinsthat folds in a certain way, what do you think

the structure of this protein, whose structureis not yet solved experimentally, should be ??

The bottom line . . . . . What is your best guess ?


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In perspectiveIn perspective

Human has 30K to 35K Genes

Number of proteins expressed in human alone canbe 10 X number of Genes

Total number of proteins expressed by all organisms

.. MANY

Number of protein structures at PDB as of May 05is 31K

There is a good chance that the protein you arelooking for is not solved !!!!


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Some Basic TheorySome Basic Theory

When is homology modeling applicable

When the sequence similarity is greater than 40 %

When there is more than one instance that supportour initial assumption. E.g. If you can find a proteinfrom different organisms with slightly different

protein sequence sharing a very similar proteinstructure (fold).


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ProcedureProcedure

Assign coordinates from SCR

Align Target Sequence to SCR sequence

Extract Sequence

Struct/Seq Alignment

Get homologous protein structure

Load Target Sequence

Generate loops

Repair the ends

Optimize structure

Determine SCR

Similar Sequence

Find homologous proteins


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Getting a Protein StructureGetting a Protein Structure

Go to protein data bank at

http://www.rcsb.org

Find a series of proteins structure thatare homologous to your protein.

Download the coordinate file (in PDBformat) to your working directory


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InsightII:InsightII:

One stop Graphical User InterfaceOne stop Graphical User Interfacefor life sciencesfor life sciences


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InsightII features cover todayInsightII features cover today

Molecular Visualization

Structure comparison Sequence alignment

Structure building

Molecular Modeling Energy Calculation

Minimization


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How to run InsightIIHow to run InsightII

Available on SGI and Linux workstations

module load insightII

start_insightII


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InsightIIInsightIIMSI logo

Viewer toolbar

Module toolbar

Frequently

Used buttons

Frequently used buttons


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LoadingLoading

moleculemolecule

Note: Molecule/GetNote: Molecule/Get


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DisplayingDisplaying

StructureStructure

Note: Molecule/DisplayNote: Molecule/Display


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HomologyHomology

Here I am

Homology toolbar

Note: Accelrys Logo/homologyNote: Accelrys Logo/homology


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ExtractingExtracting

SequenceSequence

Sequence Viewing

Window

Note: Sequences/ExtractNote: Sequences/Extract


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Manipulating Sequence Manually with the mouseManipulating Sequence Manually with the mouse

Toggle on Seq on Sequence Viewing Window

Moving Entire Sequence

Middle click on a residue and drag to move the entire sequence Creating Gap

Right click on a residue and drag to the right to create gap to theright of residue

Left click on a residue and drag to the left to create gap to the leftof residue

Moving entire Gap Right click on a gap and drag to the left to move entire gap to left Left click on a gap and drag to the right to move entire gap to right

Splitting Gap Right click on a gap and drag to the right to split gap to right Left click on a gap and drag to the left to split gap to left


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MultipleMultiple

SequenceSequenceAlignmentAlignment

Note: Alignment/Multiple SequenceNote: Alignment/Multiple Sequence

SuperimposeStructure usingSCR


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AlignedAlignedStructureStructure


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Aligned SequenceAligned Sequence

Structurally ConservedRegions (SCR)


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GetGet

SequenceSequence

Note: Sequences/GetNote: Sequences/Get


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PairwisePairwise

SequenceSequenceAlignmentAlignment

Note: Alignment/Note: Alignment/Pairwise_SequencePairwise_Sequence


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Aligned SequenceAligned Sequence


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SequenceSequenceIdentityIdentity

Note: Alignment/Note: Alignment/Percent_IdentityPercent_Identity


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Manipulating Boxes Manually with the mouseManipulating Boxes Manually with the mouse

Toggle on Box on Sequence Viewing Window

Creating Boxes

Left click on one residue of a sequence and drag to anotherresidue of another sequence to create a box Altering Boxes

Right click on left or right side of box and drag to shrink orexpand box

Moving Boxes Middle click on edge of box to move box

Freezing/Unfreezing Boxes Middle click then left click to freeze box

Repeat to unfreeze box Deleting Boxes

Middle click, hold down control on keyboard, then left clickto delete box


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Assigning SCR manuallyAssigning SCR manually


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AssigningAssigning

CoordinatesCoordinatesfrom Alignedfrom AlignedSequenceSequence

Note: Sequences/Note: Sequences/Assign_CoordsAssign_Coords


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GeneratingGenerating

LoopsLoops

Note: Loops/GenerateNote: Loops/Generate


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LoopsLoops

Note: Loops/DisplayNote: Loops/Display


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AssignAssign

CoordinatesCoordinatesto Loopsto Loops



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SettingSetting

VariableVariableto Localto LocalPDBPDB

Note: Session/Note: Session/Env_VarEnv_Var//usr/local/db/pdb/current/entriesusr/local/db/pdb/current/entries


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SearchingSearching

LoopsLoops

Note: Loops/SearchNote: Loops/Search


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End RepairEnd Repair

Note: Refine/Note: Refine/EndRepairEndRepair


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TraceTrace



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SCR andSCR and

LoopLoopRegionsRegions



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RelaxRelaxSetupSetup

Note: Refine/RelaxNote: Refine/Relax


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RelaxRelax

Note: Refine/RelaxNote: Refine/Relax


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Compare withCompare with

ExperimentalExperimentalStructureStructure(1CD2)(1CD2)


R i


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RunningRunning

FastaFasta

Note: Databases/InputNote: Databases/Input

R iR i


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RunningRunning

FastaFasta

Note: Databases/OutputNote: Databases/Output

R iR i


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RunningRunning

FastaFasta

Note: Databases/RunNote: Databases/Run

F tF t


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FastaFasta

PrintoutPrintout

F tF st


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FastaFasta

PrintoutPrintout


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Learning more about InsightIILearning more about InsightIIand its modules on your ownand its modules on your own

Pilot tutorials


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InsightII DocumentationInsightII Documentation

http://www.accelrys.com/doc/life/index.html

Username: msi

Passwd: msi-doc


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To get helpTo get help

By mail [email protected]

Web www.msi.umn.edu

Phone 612 626 0802

Appointment TBA


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Contact InformationContact Information

Yuk Sham

Computational Biology/Biochemistry Consultant

Phone: (612) 624 7427 (Walter Library)

Phone: (612) 624 0783 (VWL)Email: [email protected]

homo logy modeling

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