GEB 406 Course Instructor: Sheikh Ahmad Shah Semester: Summer 2016 Lecture - 11 Protein-Protein Interaction GEB 406 Course Instructor: Sheikh Ahmad Shah Semester: Summer 2016

Protein-Protein Interaction Proteins work together by actually binding to form multicomponent complexes that carry out specific functions. These functional units can be as simple as dimeric transcription-factor complexes or as complex as the 30-plus component systems that form ribosomes. Biochemists believe that all proteins bind to or interact with at least one other protein. The discovery that proteins in higher organisms (e.g., human and mouse) contain higher numbers of functional domains suggests that many of these proteins have multiple associations. Understanding how protein complexes work is essential to understanding how cells work as systems.

Identifying Protein-Protein Interaction In the pregenomic era, immunoprecipitation was the primary means of determining protein-protein interaction.

Identifying Protein-Protein Interaction In the post genomic era, few methods have proven especially helpful for this purpose. Yeast two hybrid (Y2H) system is one of them. In short, the Y2H method is designed to use a protein of interest as bait in order to discover proteins that physically interact with the bait protein, those proteins are termed as preys. In Y2H method, a single transcription factor is cut into two pieces called the DNA Binding Domain (DBD) and Activation Domain (AD), which stimulates the RNA polymerase to begin transcription.

Identifying Protein-Protein Interaction Fused to the DBD is the bait protein of interest (B), which cannot initiate transcription on its own. Fused to the AD the prey ORF, which can be any known or unknown protein. The prey protein of AD + ORF fused together cannot initiate transcription either. When the bait and prey proteins are produced in the same cell, they might interact; and if they do, transcription of His3 gene is initiated. Any ORF can be tested with Y2H, which means a proteome-wide survey can be performed rapidly by transforming a genomic library into cells that contain bait plasmids. In this way, every protein in a proteome can be tested individually for its potential interact with bait.

Identifying Protein-Protein Interaction