Poly-lysine supported cross-linked enzyme aggregates of penicillin G acylase and its application in synthesis of beta-lactam antibiotics

摘要

Penicillin G acylase (PGA) from Providencia rettgeri PX04 (PrPGA) was utilized to synthesize beta-lactam antibiotics. Poly-lysine supported cross-linked enzyme aggregates (PL-CLEAs) were prepared using PGA. Addition of polylysine significantly increased retention of PGA activity in CLEAs, with a decrease in the synthesis/hydrolysis (S/ H) ratio. PL-CLEAs with 0.56 mg/mL poly-lysine retained 83% of free PGA activity, and displayed a higher S/H ratio than that of the free enzyme. Both PL-CLEAs and CLEAs exhibited high pH and thermal stabilities. PL-CLEAs possessed the best stability profile, and the lowest alpha value [(k(cat)/K-m)p(s)/(k(cat)/K-m)(AD)], and was most effective at amoxicillin synthesis. A >94% yield of amoxicillin was achieved using a D-HPGME/6-APA ratio of 1.2:1 (240 mM, 200 mM), with fed-batch addition of D-HPGME. PL-CLEAs displayed excellent operational stability during amoxicillin synthesis. Over 97% of initial conversion was retained after twenty rounds of catalysis. PL-CLEAs exhibited greater potency than CLEAs in practical catalysis, permitting a higher concentration of reactants. (C) 2019 Elsevier B.V. All rights reserved.