RUI: Mechanism and Function of Subunit Interactions in Oligomeric Proteins

Gustavus Adolphus College, Saint Peter MN

Investigators

Abstract

9986278 Bell Abstract: The exquisite control of the levels of metabolites within the cell involves a combination of both long-term adaptation involving control of gene expression and short-term regulation involving either the covalent modification of an enzyme, frequently by phosphorylation, or the binding of an allosteric regulatory ligand. The effectiveness of allosteric regulation is amplified in oligomeric enzymes by subunit interactions leading to cooperative interactions between regulatory ligands or between substrate binding events and catalysis. While several allosteric models have been presented to explain the roles that allosteric effects can play in the regulation of activity, the work proposed will investigate the requirement for subunit interactions in the normal catalytic cycle of an oligomeric enzyme, referred to as "reciprocating subunits". The major aims of the work are to understand the role that subunit interactions play in the regulation [both homotropic and heterotropic] and activity of four important enzymes, Malate Dehydrogenase, Fumarase, 3-Phosphoglycerate dehydrogenase and Glutamate dehydrogenase, all of whose 3-dimensional structures are available. This work will provide direct evidence for or against the existence of a reciprocating subunit mechanism with these four enzymes. We will also test our hypothesis that reciprocating subunit mechanisms may also play an integral role in regulation of these enzymes, and will test whether our proposed mechanism where heterotropic regulators of either the K-type or the V-type effect subunit-subunit interactions to elicit their effects is appropriate for these enzymes. This work will involve the construction of a series of hetero-oligomers containing combinations of "normal" and modified sites to thoroughly test the roles that ligand induced changes across subunit interfaces play in the regular activity of these enzymes. Furthermore, working in conjunction with several crystallography labs, we will be able to obtain detailed structural information as to the mechanism of the subunit interactions involved in the activity and regulation of these enzymes.