If nature did not allow protein-protein interactions to occur inside living cells (and viruses), life as we know it would simply not exist. Molecular biologists have long struggled with developing methods to study protein-protein interactions.
Recent data presented at the Gordon Research Conference on Protein Interaction Dynamics in Galveston, TX, suggest that they soon might be able to breathe a little easier.
Eric Sundberg, Ph.D., principal scientist at Boston Biomedical Research Institute, discussed “some of the complex factors that regulate protein molecular recognition, including cooperation among amino acid residues within protein interfaces and disordered protein regions involved in protein binding.”
Dr. Sundberg’s lab uses directed evolution techniques, including phage display and yeast display, to insert random mutations that evolve proteins for better binding, all in an effort to “create model systems to study how these complex factors regulate protein-protein interactions.”
Dr. Sundberg explained that “directed evolution is just like Darwinian evolution in that you go through iterative rounds of mutation and selection. Our selection for fitness would be through tighter binding. And these directed evolutionary techniques are common in the field.”
He and his team have used phage display to alter the affinity of protein-protein complexes by mutating one protein in the complex. Using the T-cell receptor and Staphylococcal superantigen, they found that mutating one protein in a model complex increases its affinity for its partner proteins in the complex.
“The particular region of the protein that we targeted in this study is a disordered disulfide loop that is disordered in its unbound state. Also, in the wild type, it is largely disordered in the complex (bound) state.
“When we apply our evolutionary techniques, we mutate a stretch of five contiguous residues within that loop. And so, when they mutate, binding affinity increases, and then we select for variants that have higher affinity.”
By isolating and analyzing (by x-ray crystallography) various intermediate complexes along an affinity continuum, Dr. Sundberg’s team was able to conclude that the random mutations caused greater disorder-to-order transitions, which, in turn, was the reason for increased affinity in the binding region.