On methodological errors in estimating the initial velocities of enzyme-catalyzed reactions and on approaches to their correction(a case study with lactate dehydrogenase)

Abstract

A universal algorithm is proposed to compute the initial velocities of enzyme-catalyzed reactions at “zero time” from the kinetic curve for substrate consumption or product accumulation determined by direct measurements of changes in enzyme activities. The research is illustrated with a commercial lactate dehydrogenase sample and a standard biochemical test kit. The proposed approach makes it possible to virtually eliminate systematic errors in measurements, determine the initial velocity with the same accuracy regardless of the levels of enzyme activities, reduce time spent on analysis to one minute, and expand the scope of application of standard test kits for studying enzymes isolated from different sources when solving a wide range of problems. The algorithm cannot be applied to analyses of coupled enzyme reactions because there is a long initial lag phase in respective kinetic curves.