DEFINITION:Discipline that deals with studying the

INTERACTOME i.e. the interactions (and their

consequences) between & among molecules

within a cell, generally proteins.

More…

It is an example of "top-down" systems

biology. Large sets of genome-wide data are

collected, & correlations between different

molecules are inferred. From the data new

hypotheses are formulated, & tested by new

experiments, & generalizations are made for

comparable molecules.

Contd…

The field of interactomics is rapidly expanding

and developing. While no biological

interactomes have been fully characterized,

over 90% of proteins in Saccharomyces

cerevisiae have been screened & their

interactions characterized, making it the first

interactome to be nearly fully specified.

SIGNIFICANCE:Most biomolecules interact with other

molecules. These interfaces (►Interfaceome)

are highly conserved throughout evolution to

avoid undesirable interactions that lead to fatal

disorders in cells (►Misinteraction Disease

theory).

De novo METHODS:►Yeast two-hybrid system

► Tandem Affinity Purification

► Co-immunoprecipitation

► Fluorescent Resonance Energy Transfer

(FRET)

► Surface Plasmon Resonance (SPR)

► X-ray tomography, Optical fluorescence

microscopy, etc.

Yeast Two-hybrid system:

X Y

X Y

RNAPol

Protein X expressed as a fusion with DNA-binding domain

Protein Y expressed as a fusion with Transcriptional Activator

Interaction results in activation of the gene

Tandem Affinity Purification :

Aff

init

y C

olu

mn

Imm

un

ogl

ob

ulin

/ C

alm

od

ulin

Protein A CBP Protein X NH2

Protein Y

Co-immunoprecipitation :Interaction between two proteins is indicated if the

immunoprecipitation of one protein results in the co-precipitation of the other.

Fluorescent Resonance Energy

Transfer (FRET):Interaction between two proteins is indicated where

energy is transferred from an excited donor

fluorophore to a nearby acceptor fluorophore. FRET

occurs only when the two fluorophores are up to 10nm apart.

Surface Plasmon Resonance :Interaction between protein X (immobilized on metal

surface) and protein Y (free in solution) is

demonstrated by a change in the refractive index of

the surface layer. SPR has been used for detectinginteractions on protein chips.

In silico METHODS:

►Protein Structural Interactome Map

(PSIMAP) Algorithm

►Accessible Surface Area (ASA) Method

►Voronoi Diagram

►Domain Fusion (Rosetta stone)

PSIMAP Algorithm :

The Protein Structural Interactome Map, is a

database of all the structurally observed interactions

among protein domains of known three-dimensional

structures in the PDB. It can be constructed using any

reliable protein domain definition, where domains are

defined as evolutionarily conserved structural and

functional protein units.

Contd…

PSIMAP Algorithm :Most commonly used domain definitions used include

SCOP(Structural classification of Proteins), Pfam

(Protein families), CATH, FSSP, etc.

PSIMAP provides an overview of all the observed

domain-domain interactions at the protein family or

superfamily level. Domains from a multi-domain PDB

entry are empirically denoted as interacting with each

other if at least 5 residue pairs are within a 5

Angstrom distance.

Accessible Surface Area Method :The ASA method detects protein regions that are

buried to be excluded from a solvent when forming a

multimer or a complex. If more than two subunits

interact or aggregate with each other, they lose some

area that could be accessible by a solvent in the state

of free subunit or domain.

Contd…

Accessible Surface Area Method :Relative accessibilities are calculated for each amino

acid in the protein by expressing the summed residue

accessible surface as a percentage of that observed

in an ALA-X-ALA tri-peptide. Surface residues are

defined as those residues that have a relative ASA of

more than 5%. Interior residues are defined as those

residues that have a relative ASA of less than 5%.

Voronoi Diagram:The Voronoi diagram, known as Dirichlet tessellation,

has been widely used in the fields of science and

engineering. The Voronoi diagram was first

introduced as an application to the study of protein

structure by Richards et al.

Contd…

Light blue circles (atoms) are

contained in domain A and

green atoms are in domain B.

Dotted lines denote Voronoi

edges and a solid line

represents the interface-faces

between two domains. Any

polygon which is adjacent to at

least one interface-face is

called interface-cell. If a cell is

an interface-cell, then we call

the atom in the cell an

interface-atom. Interface-

atoms are slightly darker thannon-interface atoms.

Domain Fusion:Gene X Gene Y ?

Gene X Gene Y

Gene X Gene Y

Genome 1

Genome 1

Genome 2

Use Gene X as query to screen Genome 2

Probable functional association

UTILITY:Rational drug design & discovery include a

step for identifying interaction sites of lead

compound to the target molecules using

computers. Identifying & classifying protein

interaction interfaces on a large scale can help

researchers identify drug targets more

efficiently.

Target Validation

Target Identification

Hit Identification

Lead Identification

Lead Optimization

Non-clinical development &

Clinical Trials

The Drug Discovery Process

SHORTCOMINGS:• Noisy results

• Technique determines which interactions are found

• Interactomes may vary between tissues and developmental stages