Lab 4.3

Comparative Modeling

by MODELLER

Roberto Sanchez

Mount Sinai School of Medicine

New York, NY, USA

roberto.sanchez@physbio.mssm.edu

http://physbio.mssm.edu/~sanchez/

Comparative Modeling by MODELLER

• Using MODELLER

• Example 1: Modeling of BLBP

• Example 2: Protein/ligand complex model

• Example 3: Multiple template modeling

Comparative Modeling by MODELLER

• Obtaining MODELLER and related info• Using MODELLER

• Example 1: Modeling of BLBP

• Example 2: Protein/ligand complex model

• Example 3: Multiple template modeling

Obtaining MODELLER and related information

• MODELLER web pagehttp://guitar.rockefeller.edu/modeller/modeller.html

• Download Software (UNIX/Linux)

• HTML Manual

• Join Mailing List

Using MODELLER

• No GUI! • Controlled by command file (script) • Script is written in TOP language • TOP language is simple

Using MODELLER

• INPUT:• Target Sequence (PIR format)

• Template Structure (PDB format)

• TOP command file

• OUTPUT:• Target-Template Alignment

• Model in PDB format

• Other data

EXAMPLES

• Example files are in:

/PATH/examples/• Copy example files to your home directory with

the following command:

cp –R /PATH/examples .

Example 1: Modeling of BLBPINPUT

• Target: Brain lipid-binding protein (BLBP)• BLBP sequence in PIR (MODELLER) format,

file examples/ex1/blbp.seq :

>P1;blbp

sequence:blbp::::::::

VDAFCATWKLTDSQNFDEYMKALGVGFATRQVGNVTKPTVIISQEGGKVVIRTQCTFKNTEINFQLGEEFEETSIDDRNCKSVVRLDGDKLIHVQKWDGKETNCTREIKDGKMVVTLTFGDIVAVRCYEKA*

• Template search already done for you:Template: PDB file 1HMSexamples/ex1/1hms.pdb

Example 1: Modeling of BLBPSTEP 1: Align blbp and 1hms sequences TOP script for target-template alignment

File examples/ex1/align.topREAD_MODEL FILE = '1hms.pdb'

SEQUENCE_TO_ALI ALIGN_CODES = '1hms'

READ_ALIGNMENT FILE = 'blbp.seq', ALIGN_CODES = 'blbp', ADD_SEQUENCE = on

ALIGN

WRITE_ALIGNMENT FILE='blbp-1hms.ali', ALIGNMENT_FORMAT = 'PIR'

WRITE_ALIGNMENT FILE='blbp-1hms.pap', ALIGNMENT_FORMAT = 'PAP'

Run by typing mod align.top in the examples/ex1/ directory.If something goes wrong check file align.log

Example 1: Modeling of BLBPSTEP 1: Align blbp and 1hms sequences

OUTPUT

File examples/ex1/blbp-1hms.ali>P1;1hms

structureX:1hms: 1 : : 131 : :undefined:undefined:-1.00:-1.00

VDAFLGTWKLVDSKNFDDYMKSLGVGFATRQVASMTKPTTIIEKNGDILTLKTHSTFKNTEISFKLGVEFDETTA

DDRKVKSIVTLDGGKLVHLQKWDGQETTLVRELIDGKLILTLTHGTAVCTRTYEKE*

>P1;blbp

sequence:blbp: : : : : : : 0.00: 0.00

VDAFCATWKLTDSQNFDEYMKALGVGFATRQVGNVTKPTVIISQEGGKVVIRTQCTFKNTEINFQLGEEFEETSI

DDRNCKSVVRLDGDKLIHVQKWDGKETNCTREIKDGKMVVTLTFGDIVAVRCYEKA*

Example 1: Modeling of BLBPSTEP 1: Align blbp and 1hms sequences

OUTPUT

File examples/ex1/blbp-1hms.pap _aln.pos 10 20 30 40 50 60

1hms VDAFLGTWKLVDSKNFDDYMKSLGVGFATRQVASMTKPTTIIEKNGDILTLKTHSTFKNT

blbp VDAFCATWKLTDSQNFDEYMKALGVGFATRQVGNVTKPTVIISQEGGKVVIRTQCTFKNT

_consrvd **** **** ** *** *** ********** **** ** * * *****

_aln.pos 70 80 90 100 110 120

1hms EISFKLGVEFDETTADDRKVKSIVTLDGGKLVHLQKWDGQETTLVRELIDGKLILTLTHG

blbp EINFQLGEEFEETSIDDRNCKSVVRLDGDKLIHVQKWDGKETNCTREIKDGKMVVTLTFG

_consrvd ** * ** ** ** *** ** * *** ** * ***** ** ** *** *** *

_aln.pos 130

1hms TAVCTRTYEKE

blbp DIVAVRCYEKA

_consrvd * * ***

Example 1: Modeling of BLBPSTEP 2: Model the blbp structure using the

alignment from step 1. TOP script for model building

File examples/ex1/model.topINCLUDE

SET ALNFILE = 'blbp-1hms.ali'

SET KNOWNS = '1hms'

SET SEQUENCE = 'blbp'

SET STARTING_MODEL = 1

SET ENDING_MODEL = 1

CALL ROUTINE = 'model'

Run by typing mod model.top in the examples/ex1/ directory.If something goes wrong (or just for fun) check file model.log

Example 1: Modeling of BLBPSTEP 2: Model the blbp structure using the

alignment from step 1. OUTPUT

File examples/ex1/blbp.B99990001

• PDB file

• Can be viewed with Rasmol, PDBviewer, etc.

• Transfer from SGI server to NT workstation using ftp:

ftp server

cd examples/ex1/

get blbp.B99990001

Example 2: Modeling of a BLBP/Oleic Acid complex

INPUT

• BLBP – 1hms alignment from Example 1cd examples/ex2cp ../ex1/blbp-1hms.ali blbp-1hms-ola.ali

>P1;1hmsstructureX:1hms: 1 : : 133 : :undefined:undefined:-1.00:-1.00VDAFLGTWKLVDSKNFDDYMKSLGVGFATRQVASMTKPTTIIEKNGDILTLKTHSTFKNTEISFKLGVEFDETTADDRKVKSIVTLDGGKLVHLQKWDGQETTLVRELIDGKLILTLTHGTAVCTRTYEKE.*>P1;blbpsequence:blbp: : : : : : : 0.00: 0.00VDAFCATWKLTDSQNFDEYMKALGVGFATRQVGNVTKPTVIISQEGGKVVIRTQCTFKNTEINFQLGEEFEETSIDDRNCKSVVRLDGDKLIHVQKWDGKETNCTREIKDGKMVVTLTFGDIVAVRCYEKA.*

Oleic acid molecule in 1hms PDB file is residue number 133. This can be checked by RASMOL or by looking at the PDB file directly.

Example 2: Modeling of a BLBP/Oleic Acid complex

TOP script for modeling

File examples/ex2/model2.top

Run by typing mod model2.top in the examples/ex2/ directory.If something goes wrong check file model2.log

INCLUDESET ALNFILE = 'blbp-1hms-ola.ali'SET KNOWNS = '1hms'SET SEQUENCE = 'blbp'SET STARTING_MODEL = 1SET ENDING_MODEL = 1SET HETATM_IO = onCALL ROUTINE = 'model'

Example 2: Modeling of a BLBP/Oleic Acid complex.

OUTPUT

File examples/ex2/blbp.B99990001

• View ligand with rasmol:

Menu:

Display -> backbone

Colours -> structure

Command line:

RasMol> select ligand

RasMol> color green

RasMol> cpk

Example 3: Multiple template modeling INPUT

• Target: Brain lipid-binding protein (BLBP)

• BLBP sequence in PIR (MODELLER) format,file examples/ex3/blbp.seq :

• Templates: PDB files 1HMS andexamples/ex3/1hms.pdbexamples/ex3/1lid.pdb

Example 3: Multiple template modelingSTEP 1: Align blbp and template sequences

TOP script for target-template alignment

File examples/ex3/align.topSET ALIGN_CODES = '1hms' '1lid'

SET ATOM_FILES = '1hms.pdb' '1lid.pdb'

MALIGN3D

SET ADD_SEQUENCE = on, ALIGN_BLOCK = 2

READ_ALIGNMENT FILE = 'blbp.seq', ALIGN_CODES = 'blbp'

ALIGN

WRITE_ALIGNMENT FILE = 'blbp-1hms-1lid.ali'

WRITE_ALIGNMENT FILE = 'blbp-1hms-1lid.pap', ALIGNMENT_FORMAT = 'PAP'

Run by typing mod align.top in the examples/ex3/ directory.If something goes wrong check file align.log

Example 3: Multiple template modelingSTEP 2: Model the blbp structure using the

alignment from step 1. TOP script for model building

File examples/ex3/model3.topINCLUDE

SET ALNFILE = 'blbp-1hms-1lid.ali'

SET KNOWNS = '1hms' '1lid'

SET SEQUENCE = 'blbp'

SET STARTING_MODEL = 1

SET ENDING_MODEL = 1

CALL ROUTINE = 'model'

Run by typing mod model3.top in the examples/ex3/ directory.If something goes wrong check file model3.log

Model your own protein

• Template Search (BLAST)http://www.ncbi.nlm.nih.gov/blast/

• Model Evaluation (VERIFY3D)http://www.doe-mbi.ucla.edu/verify3d.html

VERIFY3D

Alignment with errors Correct Alignment