Construction of an A2aGPCR Construction of an A2aGPCR Model using Homology Model using Homology

Comparative Protein ModelingComparative Protein Modeling

Gabrielle Roberts Gabrielle Roberts

Biology Department, Long Island University, Biology Department, Long Island University, Brooklyn, NYBrooklyn, NY

Introduction Introduction

• Brief overview of protein structure Brief overview of protein structure

• GPCR GPCR

• New Age of Bioinformatics New Age of Bioinformatics

• A2a GPCR Model PredictionA2a GPCR Model Prediction

PROTEINS PROTEINS

Diverse range of bio molecules Diverse range of bio molecules which play important structural which play important structural and functional roles within cellsand functional roles within cells

Hair & Nails: KeratinKeratin

Cytoskeleton:Microtubules, intermediate filaments, Microtubules, intermediate filaments,

microfilamentsmicrofilaments Muscle Tissues:

Actin and MyosinActin and Myosin

Protein structureProtein structure • Protein is a polymer made up of Protein is a polymer made up of

amino acid monomers. amino acid monomers.

The variable R-group accounts for the presence of 20 different amino acids

Protein Structural levelsProtein Structural levels

Primary structure

Secondary structure

Tertiary structure

Quaternary structure

Polypeptide chain

Alpha helix or beta sheets

Functional protein*

Interactions between more than one polypeptide

Secondary StructureSecondary Structure: : Beta pleated Sheets Beta pleated Sheets

• Hydrogen bond between N-H and Hydrogen bond between N-H and carboxylic group on the backbone of carboxylic group on the backbone of adjacent beta strands adjacent beta strands

Parallel Anti-Parallel

Beta structural motifsBeta structural motifsBeta hairpin •Adjacent strands in anti-parallel arrangement •Connected by 2-5 amino acids

Greek key motif •4 anti parallel stands linked by hairpins

B-meander motif 2 or more anti parallel strands linked by hair pins

Psi-loop motif- 2 anti parallel stands with one in between connected to both by hydrogen bonds

Secondary StructureSecondary Structure: Alpha Helix: Alpha Helix

• Hydrogen bond between Hydrogen bond between N-H and carboxylic groups N-H and carboxylic groups on backbone on backbone

• Helical twists Helical twists

Tertiary StructureTertiary Structure

• Interaction of R groups: Interaction of R groups:

1.1. HydrophobicHydrophobic: phenylalanine, valine, : phenylalanine, valine, tryptophantryptophan

2.2. HydrophilicHydrophilic: glutamic acid, : glutamic acid, asparaginesasparagines

3.3. Salt bridgesSalt bridges: charged amino acids : charged amino acids

4.4. Disulfide bridgesDisulfide bridges

• Larger motifs Larger motifs

Serum albumin

Homo sapiens

Pilin

Neisseria gonorrhoeae

Tertiary structural domainsTertiary structural domains

• Domain is the stable part of the Domain is the stable part of the protein protein

• Has a particular function: Has a particular function:

a.a. Binding to ligand, DNA, or another Binding to ligand, DNA, or another proteinprotein

b.b. Spanning plasma membrane Spanning plasma membrane

c.c. Catalytic site Catalytic site

GPCRGPCR

• 7 transmembrane domains that bind 7 transmembrane domains that bind to molecules such as hormones, to molecules such as hormones, neurotransmittersneurotransmitters

• Molecular binding activates signal Molecular binding activates signal transduction pathways transduction pathways

New Age of BioinformaticsNew Age of Bioinformatics Allows scientists to virtually screen possible Allows scientists to virtually screen possible

drug targets to protein. drug targets to protein.

• X-ray diffraction is used to make a crystallized X-ray diffraction is used to make a crystallized structure of protein. The structure is stored in a structure of protein. The structure is stored in a Protein Data Bank.Protein Data Bank.

http://www.pdb.org/pdb/home/home.dohttp://www.pdb.org/pdb/home/home.do

• Molecular docking Molecular docking design drugs targets that fit conformation of design drugs targets that fit conformation of

proteins or have affinity for protein receptors. proteins or have affinity for protein receptors.

PROBLEMPROBLEM

• Difficult to crystallize GPCR Difficult to crystallize GPCR

SOLUTIONSOLUTION

• Use comparative homology modeling Use comparative homology modeling to construct a theoretical 3 to construct a theoretical 3 dimensional model dimensional model

“ “domains are conserved” domains are conserved”

“ “proteins from the same family have proteins from the same family have significant similarities in tertiary significant similarities in tertiary structures” structures”

OBJECTIVE OBJECTIVE

To construct a theoretical model of an To construct a theoretical model of an A2a GPCR using homology modelingA2a GPCR using homology modeling

Method Method Select protein templateSelect protein template: :

Align target sequence(A2a) with template Align target sequence(A2a) with template sequence(A2b). Altering sequence of A2a sequence(A2b). Altering sequence of A2a to match A2b. to match A2b.

Use homology modeling program Modeller Use homology modeling program Modeller to create 3D structure of A2a to create 3D structure of A2a

View protein structure using Ras win. View protein structure using Ras win.

A2b: GPCR family, PDB

Results:Results: Protein alignmentProtein alignment • Protein sequence obtained from NCBI Protein sequence obtained from NCBI

website website

A2b is 60% identical to A2a sequence

191 residues of A2a matched A2b

Results: Results: Homology ModelingHomology Modeling

ModellerModeller: computer program that : computer program that uses altered sequence and A2b PDB uses altered sequence and A2b PDB file to make a A2a model. file to make a A2a model.

“spartial restraints” and the “position of atoms in space”

A2a Theoretical Model vs. A2b ModelA2a Theoretical Model vs. A2b Model Models viewed using Ras win

Future Considerations Future Considerations

• To attempt model assessment with To attempt model assessment with molecular docking of known A2a molecular docking of known A2a ligand antagonists/ agonistsligand antagonists/ agonists

Specific pocket binding residues Specific pocket binding residues