Purification and biochemical activity of Salmonella exchange factor SopE

Abstract

This chapter discusses the purification and biochemical activity of salmonella typhimurium exchange factor SopE. Salmonella typhimurium is a bacterial enteropathogen responsible for a large percentage of bacterial food poisoning. SopE is one of the S. typhimurium effector proteins injected into host cells. SopE are thought to provide the signals necessary for recognition and injection into host cells by the type III secretion system. The chapter presents a detailed description of the preparation and of several biochemical assays to analyze the interaction of SopE with Rho guanosine 5'-triphosphate (GTPase).The chapter explains the process of overproduction and purification of recombinant SopE proteins. Preparation of Rho guanosine 5'-triphosphate GTPase proteins is also explained. The chapter discusses the filter-binding assay for SopE-mediated guanine nucleotide exchange. In the presence of native SopE , guanine nucleotide exchange rates of [ 3 H]GDP-loaded Cdc42 are enhanced greatly. The chapter presents surface plasmon resonance assay to study SopE- Cdc42 complex. The chapter concludes with a discussion on the specificity of SopE.