Protein-protein Protein-protein interactionsinteractions

Lecture series in systems biology

Department of BioinfomaticsShanghai Jiao Tong University

Woo Mao-Ying

ricket@sjtu.edu.cn

http://202.120.45.17/course/intro/ppi.htm

Outline

Why protein-protein interactions?. Experimental methods for discovering PPIs:

• Yeast-two-hybrid （酵母双杂交）• AP-MS （亲和纯化 -质谱串联）

PPIs databases:• DIP• MIPs

Computational prediction of PPIs• Phylogenetic based method （基于进化的手段）• Expression correlation based method （基于表达相关性）• STRING (EMBL)

Why protein-protein interactions (PPI)?

Gene is the basic unit of heredity. Genomes are availabe.

genome Proteome （蛋白质组） interactome

Proteins, the working molecules of a cell, carry out many biological activities

Proteins function by interacting with other proteins.

Why protein-protein interactions (PPI)?

PPIs are involved in many biological processes: Signal transduction (信号传递 )

Protein complexes or molecular machinery （蛋白复合物或分子体系） Protein carrier （蛋白的运输） Protein modifications (phosphorylation) （蛋白质的修饰） …

PPIs help to decipher the molecular mechanisms underlying the

biological functions, and enhance the approaches for drug discovery

High throughput experimental methods for discovering PPIs

Yeast-two-hybrid (Y2H ，酵母双杂交） Ito T. et al., 2001

Uetz P. et al., 2000

Affinity purification followed by mass spectrometry (AP-MS ，亲和纯化 - 质谱串联） Gavin AC et al., 2002, 2006

Ho Y. et al., 2002

Krogan NJ et al., 2006

Y2H experiments

Idea: Bait 诱饵蛋白 (prey捕获蛋白 )

protein is fused to the binding domain (activation domain).

If bait and prey proteins interact, the transcription of the reporter gene is initiated.

High throughput screening the interactions between the bait and the prey library.

In yeast nucleus

AP-MS experiments

Fuse [a TAP tag consisting of protA (IgG binding

peptides) and calmodulin binding peptide (CBP)

separated by TEV protease cleavage site] to the

target protein

After the first AP step （亲和纯化第一步） using an

IgG (免疫球蛋白 ) matrix, many contaminants are

eliminated.

In the second AP step （亲和纯化第二步） , CBP

binds tightly to calmodulin coated beads. After

washing which removes remained contaminants and

the TEV protease, the bound meterial is released

under mild condition with EGTA (乙二醇二乙醚二胺四乙酸 ).

Proteins are identified by mass spectrometry

PPIs Databases.

DIP- Database of Interacting Protein.

(http://dip.doe-mbi.ucla.edu/ )

MIPS-Munich Information center for Protein

Sequences.

(http://mips.gsf.de/ )

DIP

Protein function Protein-protein relationship Evolution of protein-protein interaction The network of interacting proteins Unknown protein-protein interaction The best interaction conditions

DIP-Statistics

Number of proteins: 20731

Number of organisms: 274

Number of interactions: 57687

Number of distinct experiments describing an interaction:

65735

Number of data sources (articles): 3915

DIP-Searching information

Find information about your protein

DIP Node (DIP:1143N)

Graph of PPIs around DIP:1143N

Nodes are proteins

Edges are PPIs

The center node is DIP:1143N

Edge width encodes the number

of independent experiments

identyfying the interaction.

Green (red) is used to draw core

(unverified) interactions.

Click on each node (edge) to

know more about the protein

(interaction).

List of interacting partners of DIP:1143N

MIPS

Services: Genomes

Databanks retrieval systems

Analysis tools

Expression analysis

Protein protein interactions MPact: the MIPS protein interaction resource on yeast.

MPPI: the MIPS Mammalian Protein-Protein Interaction Database.

Protein complexes Mammalian protein complexes at MIPS

MPact: the MIPS protein interaction resource on yeastQuery all PPIs of a yeast protein

MPact: the MIPS protein interaction resource on yeast

MPact: Interaction Visualization

MPPI: the MIPS Mammalian Protein-Protein Interaction Database

Query PPIs of a mamalian protein. You can use x-ref, for example Uniprot accession number.

Results for PPI search

In short format

Results for PPI search

In full format

Mammalian protein complexes at MIPS

Search information of complexes

Assessment of large–scale data sets of PPIs

The overlap between the individual methods is surprisingly small

The methods may not have reached saturation. Many of the methods may produce a significant

fraction of false positives. Some methods may have difficulties for certain

types of interactions

Von Mering C, et al. Nature, (2002) 417 : 399–403

Functional biases

AP-MS discovers few PPIs involved in transport and sensing Y2H detects few PPIs involved in translation. Different methods complement each other

Von Mering C, et al. Nature, (2002) 417 : 399–403

Coverage and Accuracy

Von Mering C, et al. Nature, (2002) 417 : 399–403

• Limited and biased coverage (False Negatives)

• High error rate (False Positives)

• Expensive, time-consuming and labor-intensive

Computational methods of prediction

Current approaches:

Genomic methods

Biological context methods

Structural based methods

Genomic methods

Protein a and b whose genes are close in different genomes are predicted to interact.

Protein a and b are predicted to interact if they combine (fuse) to form one protein in another organism.

Protein a and c are predicted to interact if they have similar phylogenetic profiles.

Biological context methods

Gene expression: Two protein whose genes exhibit

very similar patterns of expression across multiple

states or experiments may then be considered

candidates for functional association and posibly

direct physical interaction.

GO annotations: two interacting proteins likely have

the same GO term annotations.

Machine learning techniques are adopted for PPI

classification by intergrating all known information.

STRING: Search Tool for the Retrieval of Interacting Genes/Proteins

A database of known and predicted protein interactions Direct (physical) and indirect (functional) associations The database currently covers 2,483,276 proteins from 630

organisms Derived from these sources:

Supported by

Searching information

Query infomation via protein names or protein sequences.

Graph of PPIs

Nodes are proteins

Lines with color is an evidence of

interaction between two proteins.

The color encodes the method

used to detect the interaction.

Click on each node to get the

information of the corresponding

protein.

Click on each edge to get

information of the interaction

between two proteins.

List of predicted partners

Partners with discription and confidence score. Choose different types of views to see more detail

Neighborhood View

The red block is the queried protein and others are its neighbors in organisms. Click on the blocks to obtain the information about corresponding proteins.

The close organisms show the similar protein neighborhood patterns. Help to find out the close genes/proteins in genomic region.

Occurence Views

Represents phylogenetic profiles of proteins. Color of the boxes indicates the sequence similarity between the proteins and

their homologus protein in the organisms. The size of box shows how many members in the family representing the

reported sequence similarity. Click on each box to see the sequence alignment.

Gene Fusion View

This view shows the individual gene fusion events per species Two different colored boxes next to each other indicate a fusion

event. Hovering above a region in a gene gives the gene name; clicking on

a gene gives more detailed information

References

Ito T et.al: A comprehensive two-hybrid analysis to explore the yeast protein interactome. Proc. Natl Acad. Sci. USA 2001, 98:4569-4574.

Uetz P et. al: A comprehensive analysis protein-protein interactions in Saccharomyces cerevisiae. Nature 2000, 403:623-627.

Gavin AC et.al: Functional organization of the yeast proteome by systematic analysis of protein complexes. Nature 2002, 415:141-147.

Gavin AC et.al: Proteome survey reveals modularity of the yeast cell machinery. Nature 2006, 440:631-636.

Ho Y et.al: Systematic identification of protein complexes in Saccharomyces cerevisiae by mass spectrometry. Nature 2002, 415:180-183.

Von Mering C et.al: Comparative assessment of large-scale data sets of protein-protein interactions. Nature 2002, 417:399-403.

Thank you for your attention