Homology

Contents

Release 2005, March 2005

1. Introduction

What is Homology?

Hardware and Installation

How to Invoke Homology

Program Environment

Homology and Insight II

Saving Homology Information with Insight II

Command Logging and Restarting

Homology and Discover

Operations

2. Theory

Background

Homology Model Building

Searching Sequence Databases With the FASTA Program

ktup Value

Scoring Local Regions

Joining Regions

Optimizing the Sequence Matching

Explicit Statistical Estimation

Determining Structurally Conserved Regions (SCRs)

Manual Determination of Structurally Conserved Regions

Automatic Determination of Structurally Conserved Regions

Automatic Sequence Alignment Methods

Needleman and Wunsch Algorithm for Pairwise Alignment

Accelrys' Pairwise Alignment Procedure

Accelrys' Automatic Multiple Sequence Alignment

align123

Scoring Matrices

Superimposing structures based on sequence alignment

Multiple Structure Alignment

Simultaneous Superposition of Structures

Assignment of Coordinates Within a Conserved Region

Assignment of Coordinates for Loop or Variable Regions

Search Loops Command

Generate Loops Command

Side Chain Conformational Searches Using Rotamer Libraries

Refinement of the Model Using Molecular Mechanics

The Potential Energy Equation

Energy Minimization

Energy Constraints

Molecular Dynamics

Secondary Structure Prediction

The Chou-Fasman Method

The GOR II Method

Hydrophobicity Profiles

Solvent Accessible Surfaces

Definition of Solvent Accessible Surface Area

Significance of Solvent Accessible Surface Area

Solvation Module versus ProStat

Solvent Accessible Surface Area for Protein Structure Validation

Rules for Protein Validation

3. Implementation

Sequence Window

Sequence Display

Controls

Sequence Boxes

Sequence Gaps

Manipulating the Sequence Display

Scrolling Modes

Seq Mode

Box Mode

4. Command summary

Modules

Pulldowns

Commands

Sequences pulldown

Boxes pulldown

Loops pulldown

Residue pulldown

Databases pulldown

Background_Job pulldown

Alignment pulldown

By_Residue pulldown

Refine pulldown

Profiles_3D pulldown

ProStat pulldown

Modeler pulldown

Seqfold pulldown

5. Methodology

Step 1: Determine Which Proteins Are Related to the Model Protein

Generating a Sequence Database

Sequence Database Searching

Motif searching

Step 2: Determining Structurally Conserved Regions (SCRs)

Automatic Determination of SCRs

Specifying the Initial Search Zone

Optimizing the Automatic Search for SCRs

Subsets

Automatic Superimposing of Structures

Characteristics of m-boxes

Handling of Existing Boxes

Interrupting the Search

Manual Determination of SCRs

Finding Pairwise SCRs

Criteria for Evaluating Manually-Determined SCRs

Summarizing the Manually-Determined SCRs

Superimposing Reference Proteins Using Manually-Determined SCRs

Superimposing structures

Multiple Sequence Alignment as an Alternative to the Manual Method

Step 3: Sequence Alignment

Choosing a Scoring Matrix

Automatic Sequence Alignment without SCRs

Automatic Sequence Alignment with Automatically-Determined SCRs

Automatic Sequence Alignment with Manually-Determined SCRs

Pairwise Manual Sequence Alignment

Multiple Sequence Alignment

Specifying the Initial Search Zone

Specifying a Mandatory Sequence

Automatic Calculation of Pairwise Threshold

Statistical Significance and Alternate Sequence Coloring

Characteristics of m-boxes

Subsets

Automatic Superimposing of Structures

Adjusting the Sensitivity and Selectivity of the Search

Handling of Existing Boxes

Interrupting the Search

Excessive Calculation Time

Single_Search Mode

Manual Mode

Step 4: Assigning Coordinates Within the SCRs

Step 5: Building Loop or Variable Regions (VRs)

Searching for and Displaying Loops

Generating and Displaying Loops

Building Coordinates for the VRs

Step 6: Conformational Search for Side Chains Using Rotamers

Step 7: Refining the Structure with Discover or CHARMm

Running Discover or CHARMm with Homology-Built Model Structures

End Repair

Splice Repair

Energy Minimization

Molecular Dynamics

Step 8: Validating Results

Structure Checking

Residue Dihedral Angles

Secondary Structure Classification

Algorithmic Implementation

United Atom Models

Setting Atomic Radii

Definition of Computed Surface Areas and their Significance

Total Surface Area

Relative Surface Area and the Tripeptide Model

Polar and Apolar Surface Area

Limitations in Implementation

Conclusion

6. Tutorial

Introduction

Hardcopy and Pilot online tutorials

Hardcopy lessons

Lesson 4a: Finding structurally conserved regions

Lesson 4b: Building SCRs and loops

Lesson 10: Finding alternative multiple sequence alignments

A. References

B. File Formats

Introduction

Amino Acid Scoring Matrices

User Scoring Matrix Files

Sequence Alignment Command

Input Databases Command

Get Sequence, Alignment and Databases commands

Secondary structure file

C. Glossary

D. seq_extract Utility

Output

Results Displayed on the Screen

Output Files

Execution Options

E. Matrices

Sequence Alignment Matrices

Identity Matrix

Codon Substitution Matrix

Dayhoff Evolutionary Mutation Matrix

Hydrophobicity Matrix

Input Databases Command Matrices

Identity Matrix

Codon Substitution Matrix

Dayhoff Evolutionary Mutation Matrix

Hydrophobicity Matrix

F. Hydrophobicity Scale Values

Amino Acid Values

Threshold Values

G. Sequence Databases

Protein Sequence Databases

NBRF

Swiss-Prot

DNA Sequence Databases

GenBank

EMBL Data Library

H. align123 Standalone

HELP 1: General help for align123

HELP 2: Help for multiple alignments

HELP 3: Help for pairwise alignment parameters

HELP 4: Help for multiple alignment parameters

HELP A: Help for protein gap parameters.

HELP 5: Help for output format options.

HELP 6: Help for profile and structure alignments

HELP B: Help for secondary structure / gap penalty masks

HELP C: Help for secondary structure / gap penalty mask output options

HELP 7: Help for phylogenetic trees

HELP 8: Help for choosing a weight matrix

HELP 9: Help for command line parameters DATA (sequences)

HELP 10: Help for tree output format options