The enhancement of the stability of penicillin acylase at high temperature and in organic solvents is essential to its industrial applications.This research work started with the production of recombinant expressed in E.coliTop10F′/pGEMKTTacPGATag.The crude penicillin acylase was purified using metal chelate chromatography

the penicillin acylase product obtained was of purity 23200 U·L-1.Encapsulation of penicillin acylase into nanogel was accomplished using the aqueous twostep in situ polymerization method

in which the penicillin acylase firstly reacted with N-acryloxysuccinimide so to generate vinyl groups on its surface

and then polymerized in aqueous medium using acrylamide as monomer and ammonium persulfate as initiator.The yield of polymerization reaction was 90%

and the diameter of penicillin acylase nanogel obtained was 12—16 nm. The penicillin acylase nanogel has shown an identical fluorescence emission spectrum compared to its native counterpart.Moreover

the encapsulation into nanogel led to a substantial enhancement of enzyme stability at high temperature and in polar solvents

as shown by the extension of t1/2 by 75fold at 60℃

and 15 and 26fold in the presence of 40%(vol)methanol and ethanol

respectively

compared to its native counterpart.