Conformational Ensembles from Experimental Data and Computer Simulations

Conformational Ensembles from Experimental Data

and Computer Simulations

Monday Speaker Abstracts

Systematic Perturbation of a Fundamental Biological Switch

Tanja Kortemme

UCSF, San Francisco, CA, USA.

Cellular protein-protein interactions can be highly interconnected. Because of this complexity, it

is often difficult to extract quantitative information on how each interaction contributes to

distinct or overlapping cellular functions, and, moreover, how changes to individual interactions

result in altered function or disease. We are developing an experimental platform for studying

perturbations to multi-functional network “hub” proteins by combining high-throughput in vivo

genetic interaction screening technology (Epistatic MiniArray Profile (E-MAP)) with mass-

spectrometry and biophysical assays. Our case study protein is the highly-conserved multi-

functional Gsp1/Ran GTPase switch that controls key eukaryotic processes. The approach first

engineers defined perturbations to Gsp1/Ran protein-protein interactions by amino acid point

mutations (“edge perturbations”). The second step determines the functional effects of these

perturbations at the cellular and organism level in the model

S. cerevisiae

. We find that E-MAPs

have a resolution that enables us to identify quantitative functional differences in vivo between

individual point mutations, even those between different amino acid substitutions of the same

residue. Our analysis reveals several classes of observed phenotypes that could be explained by

the underlying biophysical perturbations of the on/off balance of the fundamental GTPase switch

and considerable allosteric effects in the system.