Protein-protein Protein-protein interactionsinteractions
Lecture series in systems biology
Department of BioinfomaticsShanghai Jiao Tong University
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
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