SUBSTRATE SPECIFICITY OF ВACILLUS THURINGIENSIS IМV В-7324 PEPTIDASE 2

Abstract

Ai m. Investigation of substrate specificity of the purified peptidase 2 from Вacillus thuringiensis IМV В-7324. Methods. To study the substrate specificity of the peptidase 2 we used protein: elastin, fibrin, fibrinogen, collagen and casein. Determination of the optimal ratio of the enzymesubstrate was carried out a two-level factorial design. Substrate affinity (Michaelis constant, Km) of enzyme was established by the double reciprocal coordinates in the Lineweaver-Burk (1/ν0 - 1 / [S]). For calculation and graphic presentation of the results obtained by two factorial experiments there were used Box-Wilson method and computer program STATISTICA 8.0. Results. The study of hydrolysis of native protein substrates by В. thuringiensis IМV В-7324 peptidase 2 showed that the enzyme is able to cleave the fibrin, fibrinogen, casein and collagen. For effective hydrolysis of proteins according to the results of full two-way experience (CFE) was determined the optimal correlation of enzyme concentration and substrates concentration. So 1 mg of peptidase 2 is able to cleave 54 mg of fibrin, 67 mg of fibrinogen, 25 mg of casein and 49 mg of collagen. It was shown that the examine enzyme is more affinity to the fibrin and casein for that the Michaelis constant Km is 1.1 and 1.2 mg, respectively. Conclusions. The peptidase 2 of B. thuringiensis IМV В-7324 is characterized by higher specificity to the fibrin and fibrinogen than to casein and collagen, but more affinity to fibrin and casein.