Showing papers on "Penicillin amidase published in 1980"

Stabilization and Reactivation of Enzymes

Abstract: Prevention of the denaturation of enzymes is frequently indispensable to their technological application. This is exemplified by the following five operational factors of importance in the practical use of enzymes: 1. Enzymes isolated from their in Vivo environment usually become labile, and their lifetime sometimes does not exceed minutes (1–4). For example, the stability of immobilized aspartase proved insufficient for technological use of this enzyme (5). 2. For many processes, elevated temperature is desirable. The rates of chemical reactions, including enzymatic, increase with temperature; so that at higher temperatures, the required conversion of the substrate will be achieved in a shorter time or with a smaller amount of immobilized enzyme. This is very important for technological processes where the cost of the enzyme is a limiting factor, e. g., glucoamylase (6). Also, higher temperatures allow germ-free conditions to be maintained (7), which are indispensable for, say, the food industry (8,9). On the other hand, denaturation of biocatalysts intensifies (1–3) at elevated temperatures. 3. Reaction equilibrium may be such that the required products are obtained only if the reaction is carried out in an aqueous-organic mixture with a high proportion of the organic component. Under such conditions enzymes as a rule lose their catalytic activity or specificity (2,10; see also references in 11). 4. Sometimes the pH optimum of an enzymatic reaction and the pH range within which the enzyme is stable do not coincide. A good example is synthesis of penicillin antibiotics under the action of penicillin amidase; equilibrium in the synthesis is shifted toward the product in an acidic medium, where the enzyme is rather unstable (12). 5. It may not be convenient to use an enzyme immediately upon harvesting or isolation. Hence there is a need for stabilization against inactivation during prolonged storage (13).

Properties of Penicillin Amidohydrolase Immobilized on Nylon Fiber

Abstract: Penicillin amidohydrolase was partially purified from the fermented broth of Bacillus megaterium, and was immobilized on nylon fiber. The surface area of nylon fiber was increased by roughening it with fine sand and activated by acid treatment. The free amino groups on the nylon fiber exposed by such treatment were then utilized to immobilize the penicillin amidase. Enzymatic properties of penicillin amidohydrolase immobilized on the nylon fiber by covalent bonding and cross linking with glutaraldehyde were studied and compared with those of soluble enzyme. The optimal pH and temperature profile of immobilized enzyme showed only slightly broader peaks, and the values of kinetic constants, , , and , of the immobilized enzyme are only slightly greater than those of the soluble enzyme. These results suggest that the mass transfer effect on the reaction rate for the penicillin amidase immobilized on nylon fiber is not so significant as the enzyme immobilized on some other support material like bentonite. The experimental results of batch reaction agreed well with the results of computer simulation for both the immobilized and soluble enzyme systems, confirming the validity of the rate equation derived which was based on the combined double inhibition by two reaction products.

[Penicillin amidase from E. coli. The kinetic and equilibrium parameters of the enzymatic hydrolysis of 7-phenylacetamidodesacetoxycephalosporanic acid catalyzed by an immobile enzyme].

Abstract: The kinetics of 7-phenylacetamidodesacetoxycephalosporanic acid (7-PADCA) catalyzed by immobilized penicillinamidase was studied The kinetic and equilibrium parameters of the reaction were determined by analysis of the kinetic curves of the reaction product accumulation Inhibition of the enzymatic reaction by the substrate and hydrolysis products was studied It was found that the Michaelis complex completely lost its activity after attachment of the substrate second molecule to it The values of the Michaelis constants, catalytic constant and constants of inhibition by the substrate and reaction products were determined: Km = (93 +/- 11) 10(-5) M, kcat = (65 +/- 5) c-1, Ks = (14 +/- 01) 10(-2) M, K1 (FAA) = (25 +/- 03) 10(-4) M, K1 (7-ADCA) = (14 +/- 01) 10(-1) M The diffusion effect in the kinetic reaction catalyzed by immobilized penicillinamidase is discussed The values of the Thiele modulus and the actual value of Km were calculated