title: Efficient cascade synthesis of ampicillin from penicillin G potassium salt using wild and mutant penicillin G acylase from Alcaligenes faecalis.
abstract: To avoid isolation and purification of the intermediate 6-aminopenicillanic acid (6-APA), a two-enzyme two-step cascade synthesis of ampicillin from penicillin G was established. In purely aqueous medium, penicillin G hydrolysis and ampicillin synthesis were catalyzed by immobilized wild-type and mutagenized penicillin G acylases from Alcaligenes faecalis (Af PGA), respectively (Fig. 1). The βF24 G mutant Af PGA (the 24th Phenylalanine of the β-subunit was replaced by Glycine) was employed for its superior performance in enzymatic synthesis of ampicillin. By optimizing the reaction conditions, including enzyme loading, temperature, initial pH and D-PGME/6-APA ratio, the conversion of the second step of ampicillin synthesis reached approximately 90% in 240 min and less than 1.7 mole D-PGME were required to produce 1 mole ampicillin. Overall, in a 285 min continuous two-step procedure, an ampicillin yield of 87% was achieved, demonstrating the possibility of improving the cascade synthesis of ampicillin by mutagenized PGA, providing an economically efficient and environmentally benign procedure for semi-synthetic penicillins antibiotics synthesis.
authors.affiliation: School of life science, Hunan University of Science and Technology, Xiangtan 411201 Hunan, PR China; State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, PR China.
authors.forename: Senwen
authors.initials: S
authors.lastname: Deng
authors.affiliation: State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, PR China.
authors.forename: Xiaoqiang
authors.initials: X
authors.lastname: Ma
authors.affiliation: Enzyme and Fermentation Technology Laboratory, College of Light Industry Science and Engineering, Nanjing Forestry University, Nanjing 210037, PR China. Electronic address: ezhsu@njfu.edu.cn.
authors.forename: Erzheng
authors.initials: E
authors.lastname: Su
authors.affiliation: State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, PR China. Electronic address: dzhwei@ecust.edu.cn.
authors.forename: Dongzhi
authors.initials: D
authors.lastname: Wei
mesh_terms: D042481:Alcaligenes faecalis / Q000201:enzymology* / Q000378:metabolism; D000667:Ampicillin / Q000302:isolation & purification / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D004800:Enzymes, Immobilized / Q000235:genetics / Q000378:metabolism; D006868:Hydrolysis; D009154:Mutation; D010399:Penicillin Amidase / Q000235:genetics / Q000378:metabolism*; D010400:Penicillin G / Q000737:chemistry*; D015202:Protein Engineering / Q000379:methods
chemical_list: D001426:Bacterial Proteins; D004800:Enzymes, Immobilized; D000667:Ampicillin; D010399:Penicillin Amidase; D010400:Penicillin G
keywords: Ampicillin; Cascade process; Enzyme catalysis; Penicillin G acylase; Protein engineering