Application of response surface methodology to the assay of gamma-glutamyltransferase.

Abstract

Response surface methodology (RSM) offers an empirical approach to the study of clinical enzyme assays. Variables such as pH, which are difficult to characterize by using theoretical enzyme kinetics, are easily included in RSM formulations. In this investigation, we studied with RSM the change in the measured activity of gamma-glutamy-transferase (EC 2.3.2.2) as a function of changes in concentrations of donor (gamma-glutamyl-3-carboxy-4-nitroanilide) acceptor (glycylglycine), and pH. The study defined large ranges for these variables over which maximum enzyme activity is obtained: donor 6.6 to 10.2 mmol/L, acceptor 129 to 250 mmol/L, and pH 7.8 to 8.5. The RSM regression polynomial was as accurate as a previously determined enzyme kinetic equation for predicting the transferase activity from given reagent substrate concentrations. Although not yielding a mechanistic understanding of an enzyme assay, RSM studies do produce an operational understanding of how an assay functions.