Electrophoretic demonstration of specific enzyme-substrate complex between pepsin and serum albumin.

The kinetics of enzymatic reactions are usually interpreted in terms of the Michaelis-Menton (1) reaction scheme which postulates formation of an intermediate enzyme-substrate complex. Although the concept of an enzyme-substrate complex has long been of great importance to our understanding of enzymatic reactions, only in relatively recent times has the existence of such a complex been directly demonstrated for any enzyme and then only with low molecular weight substrates. Thus, Keilin and Mann (2), Stern (3), and Chance (4) have spectrophotometrically shown the existence of complexes of peroxides with catalase and peroxidases, and Doherty and Vaslow (5) have used the method of dialysis equilibrium to show a complex between acetyl-3,5-dibromo-n-tyrosine and chymotrypsin. This communication describes an electrophoretic demonstration of a complex between bovine serum albumin and pepsin during the course of the enzymatic hydrolysis of the former. Several independent lines of evidence are presented that this complex is the specific Michaelis-Menten complex capable of being activated to give rise to reaction products.