STABILIZATION OF ESCHERICHIA-COLI PENICILLIN-G ACYLASE AGAINST THERMAL INACTIVATION BY CROSS-LINKING WITH DEXTRAN DIALDEHYDE POLYMERS

The thermostabilization of penicillin G acylase (PGA) obtained from a mutant of Escherichia coli ATCC 11105 by cross-linking with dextran di aldehyde molecules, at a molecular mass of 11 500, 37 700 and 71 000 D a, was studied. The thermal inactivation mechanisms of the native and modified PGA were both considered to obey first-order inactivation kin etics during prolonged heat treatment, forming fully active but temper ature-sensitive transient states. The highest enhancement to the therm ostability of PGA was obtained using dextran-71000-dialdehyde modifica tion, as a nearly ninefold increase at temperatures above 50 degrees C . The modification of PGA by dextran-11500-dialdehyde resulted in a co nsiderable reduction of the V-m and K-m parameters of the enzyme. Howe ver, other dextran dialdehyde derivatives used for modification did no t cause a meaningful change in either V-m and K-m. Modification by dex tran dialdehyde derivatives did not result in significant change to ei ther the optimal temperature or the activation energy of PGA. All modi fied PGA preparations showed lower inactivation rate constants but hig her half-lives for inactivation than those of the native PGA at all te mperatures studied. As indicated by the half-life times and k(i) value s, dextran 71000-dialdehyde was found to be more effective at cross-li nking in the thermo-stabilization of PGA than any other agent studied in this work.