Enzyme kinetics: Teaching using polyphenoloxidase with a practical science activity

Abstract

Enzymology is one of the fundamental areas of biochemistry and involves the study of the structure, kinetics, and regulation of enzyme activity. Research in this area is often conducted with purified enzymes and extrapolated to in vivo conditions. The specificity constant, Km (Michaelis-Menten constant), Vmax (catalytic power), IC(50) and Ki (inhibition constant) expresses the efficiency of an enzyme as a catalyst. These parameters are usually determined for enzyme catalysis and inhibition study. In this work we propose a classroom experiment for its determination using polyphenol oxidase (P PO) enzyme from banana. In addition, inhibition experiment was carried out by L-cysteine and p-aminobenzene sulfonamide which is classical PPO inhibitors. The Km and Vmax values of banana PPO towards catechol and 4-methyl catechol were determined by Lineweaver Burk method. Although there were a few variations in the kinetic parameters results among students, the students generally carried out the program successfully by correctly identifying inhibition type. The inquiry-based problem solving approach of this curriculum facilitates an understanding of the basic concepts of biochemistry and its application.