Protein-protein interactionsThree types of interactionsStable complexesYeast-two-hybrid screensSlide 5Slide 6Computational methodsSlide 8Slide 9Slide 10Slide 11Slide 12Protein chipsSlide 14Slide 15Protein-protein interactionsThree types of interactions•Interactions between domains in multidomain proteins•Interactions between the domains in stable complexes•Interactions between proteins in transient interactionsStable complexes•The proteins in stable complexes tend to be more closely co-expressed and more closely conserved compared to proteins in transient interactions and monomeric proteinsYeast-two-hybrid screens•Uses the transcription of a reporter gene driven by the Gal4 transcription factor to monitor whether or not two proteins are interactingYeast-two-hybrid screensDBD = transcription factorAD = activation domainRNApolBAGal4-ADGal4-DBDYeast-two-hybrid screensDBD = transcription factorAD = activation domainRNApolBAGal4-ADGal4-DBDReporter geneComputational methods•Conservation of gene order–In prokaryotes, genes are organized into operons of co-regulated and co-expressed groups of genes–Genes that are consistently part of the same operon across different, distantly related genomes are likely to be part of the same protein complex or functional process across all species–They have been selected to remain as a co-regulated unit throughout the extensive shuffling of gene order that takes place in prokaryote genomesComputational methodsComputational methods•Gene fusions–Look for cases across a set of genomes where two or more orthologs are part of the same gene in one genome, presumably as a result of gene fusion–The prediction is that the orthologs that are separate genes in other genomes interact with each other–Fused proteins are not only co-regulated, as in the conservation of gene order, but also permanently co-localized in the cell–This method is limited to certain classes of protein-protein interactions: •members of the same stable complex•proteins in the same metabolic pathwayComputational methodsyeastE. coliComputational methods•Phylogenetic methods–Relies on the detection of homologs in a set of genomes–If the pattern of homolog presence or absence is the same in a group of proteins, then these proteins are clustered together as belonging to the same functional classComputational methodsGene 1 Gene 2 Gene 3 Gene 4 Gene 5 Gene 6Genome 1Genome 2Genome 3Genome 4Predicted to interactProtein chips•Miniature devices on which proteins, or specific capture agents that interact with proteins, are arrayed•Protein chips can act to –separate proteins on the basis of specific affinity –characterize proteins if the capture agent is highly specific (e.g., antibodies)•Advantage: ultra-high throughput analysisProtein chips•Antibody chips–Arrayed antibodies used to detect and quantify specific proteins•Antigen chips–Arrayed protein antigens used to detect and quantify antibodies•Universal protein arrays–Any kind of proteins arrayed to detect or characterize protein-protein and protein-ligand interactions•Protein capture chips–Arrayed non-protein capture agents•Solution arrays–Coded microspheres or barcoded nanoparticles next generationProtein
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