On Methodological Errors in Estimating the Initial Velocities of Enzyme-Catalyzed Reactions and Approaches to Their Correction: A Case Study with Lactate Dehydrogenase

Abstract

A universal algorithm for calculating the initial velocities of enzymatic reactions at the “zero” moment of time according to the kinetic curve of substrate consumption or product accumulation during direct kinetic measurements of enzyme activity is presented. The study was performed using the example of a commercial lactate dehydrogenase preparation and a standard biochemical set of reagents. The proposed approach makes it possible to practically eliminate systematic measurement error, to determine the initial reaction velocity with equal accuracy regardless of the level of enzyme activity, to reduces the analysis time to 1 min, and to expand the scope of application of standard reagent kits for the study of samples of enzymes of different origin in solving a wide range of tasks. The algorithm cannot be applied when using conjugated enzyme systems in an analytical procedure, since in this case there is a long initial lag phase on the kinetic curves.