title: Efficient whole-cell biocatalyst for Neu5Ac production by manipulating synthetic, degradation and transmembrane pathways.
abstract: OBJECTIVE
  To develop a strategy for producing N-acetyl-D-neuraminic acid (Neu5Ac), which is often synthesized from exogenous N-acetylglucosamine (GlcNAc) and pyruvate, but without using pyruvate.
  
  
  RESULTS
  An efficient three-module whole-cell biocatalyst strategy for Neu5Ac production by utilizing intracellular phosphoenolpyruvate was established. In module I, the synthetic pathway was constructed by coexpressing GlcNAc 2-epimerase from Anabaena sp. CH1 and Neu5Ac synthase from Campylobacter jejuni in Escherichia coli. In module II, the Neu5Ac degradation pathway of E. coli was knocked out, resulting in 2.6 ± 0.06 g Neu5Ac l-1after 72 h in Erlenmeyer flasks. In module III, the transmembrane mode of GlcNAc was modified by disruption of GlcNAc-specific phosphotransferase system and Neu5Ac now reached 3.7 ± 0.04 g l-1. In scale-up catalysis with a 1 l fermenter, the final Neu5Ac yield was 7.2 ± 0.08 g l-1.
  
  
  CONCLUSIONS
  A three-module whole-cell biocatalyst strategy by manipulating synthetic, degradation and transmembrane pathways in E. coli was an economical method for Neu5Ac production.
authors.affiliation: The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, 214122, China.
authors.forename: Deqiang
authors.initials: D
authors.lastname: Zhu
authors.affiliation: The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, 214122, China. xbzhan@yahoo.com.
authors.forename: Xiaobei
authors.initials: X
authors.lastname: Zhan
authors.affiliation: The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, 214122, China.
authors.forename: Jianrong
authors.initials: J
authors.lastname: Wu
authors.affiliation: The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, 214122, China.
authors.forename: Minjie
authors.initials: M
authors.lastname: Gao
authors.affiliation: The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, 214122, China.
authors.forename: Zhongsheng
authors.initials: Z
authors.lastname: Zhao
mesh_terms: D000117:Acetylglucosamine / Q000378:metabolism; D017033:Anabaena / Q000378:metabolism; D055162:Biocatalysis; D016123:Campylobacter jejuni / Q000378:metabolism; D002238:Carbohydrate Epimerases / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D019158:N-Acetylneuraminic Acid / Q000378:metabolism*; D010728:Phosphoenolpyruvate / Q000378:metabolism; D019289:Pyruvic Acid / Q000378:metabolism
chemical_list: D002352:Carrier Proteins; D010728:Phosphoenolpyruvate; D019289:Pyruvic Acid; D002238:Carbohydrate Epimerases; C024519:N-acyl-D-glucosamine 2-epimerase; D019158:N-Acetylneuraminic Acid; D000117:Acetylglucosamine
keywords: Coexpressing vector; GlcNAc-specific PTS; Neu5Ac; NeuNAc synthase; Phosphoenolpyruvate; Whole-cell biocatalyst