Protein-protein interactions

Three types of interactions

• Interactions between domains in multidomain proteins

• Interactions between the domains in stable complexes

• Interactions between proteins in transient interactions

Stable 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 proteins

Yeast-two-hybrid screens

• Uses the transcription of a reporter gene driven by the Gal4 transcription factor to monitor whether or not two proteins are interacting

Yeast-two-hybrid screens

DBD = transcription factorAD = activation domain

RNApol

BA

Gal4-AD

Gal4-DBD

Yeast-two-hybrid screens

DBD = transcription factorAD = activation domain

RNApol

BA

Gal4-AD

Gal4-DBD

Reporter gene

Computational 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 genomes

Computational methods

Computational 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 pathway

Computational methods

yeast

E. coli

Computational 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 class

Computational methods

Gene 1 Gene 2 Gene 3 Gene 4 Gene 5 Gene 6

Genome 1

Genome 2

Genome 3

Genome 4

Predicted to interact

Protein 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 analysis

Protein 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

generation

Protein chips