Assessing Protein Interactions for Clustering of Mitochondrial Urea Cycle Enzymes.

Abstract

Enzyme clustering is a phenomenon that involves partitioning of proteins that function together in a common subcellular or sub-organellar compartment. Traditional genetic, biochemical, and biophysical approaches for studying protein-protein interactions in complexes with defined stoichiometry yield inconclusive results when applied to clustered proteins. This chapter describes a combination of approaches to study clustered proteins including co-immunoprecipitation, biochemical co-localization in purified mitochondria, and super resolution imaging of endogenous proteins in situ. These approaches can be used to study interactions among proteins that form clusters. We will illustrate this approach by using the urea cycle enzymes that localize in the mitochondrial matrix, and form clusters at the inner mitochondrial membrane.