用电子束辐射法制备固定化乳酸脱氢酶

PREPARATION OF IMMOBILIZED LACTATE DEHYDROGENASE BY ELECTRON-BEAM IRRADIATION

前言固定化酶的制备,近年来已有许多进展.固定化技术之一的凝胶格子包埋法,尤其受到重视,因为它与其它方法不同,酶分子本身在固定化过程中几乎完全不发生结合或偶联反应,而是被包埋在凝胶的微细格子里.这样,它可能对酶蛋白的构象。

A new and simple method for immobilization of lactate dehydrogenase by radiopolymerization and crosslinking of acrylamide or polyvinyl alcohol (PVA) is reported,when a mixture of the enzyme solution and acrylamide monomer or polyvinyl alcohol polymer was irradiated under an electron-beam at -20℃,the enzyme was entrapped in the resulting gel.Thus,two types of immobilized enzyme,polyacrylamide-lactate dehydrogenase(PAA-LDH) and polyvinyl alcohol-lactate dehydrogenase (PVA-LDH) were obtained.It was found that lactate dehydrogenase was very stable against about 6 Mrad of irradiation dose at Iow temperatures. Each type had its own merits concerning activity,mechanical strength and preparation steps. The general properties of the prepared and native lactate dehydrogenase were compared.The optimum activity temperature of PAA-LDH gel was higher than that of the native enzyme,but that of PVA-LDH gel was lower. The optimum pH for PAA-LDH gel was 10.0,similar to that of the native enzyme,and for PVA-LDH gel was 10.4. The apparent activities of these enzyme gels were 12-56 per cent of the initial activities.No activity decrease was observed when the enzyme reaction was repeated 18 times. After eight months in a refrigerator at 4℃,the activities of lyophylized enzyme gels still possessed their original vigour.