Differences in the Average Single Molecule Activities of E. coli β-Galactosidase: Effect of Source, Enzyme Molecule Age and Temperature of Induction

Abstract

Using a capillary electrophoresis–based method, single enzyme molecule assays were performed on E. coli β-galactosidase from three different sets of samples. The first set consisted of lysates of induced cells from five different strains of the bacteria, as well as two different commercial preparations of the enzyme. These samples were found to have substantially different distributions of single molecule activities. For the second set of samples, β-galactosidase expression was induced for 1.5 hr, followed by further incubation where expression was repressed. Assays were performed on the lysates of the preinduction and on the lysates from aliquots taken set times postinduction. The recently induced enzyme had a 25% higher average single molecule activity than the basally expressed enzyme. This average activity returned to the basal value 3.5 hr postinduction and remained unchanged thereafter. Finally, β-galactosidase was induced at 26 and 42°C. The enzyme was assayed before and after partial thermal denaturation. The samples were found to be indistinguishable with respect to their average single molecule activities.