title: Engineering of an N-acetylneuraminic acid synthetic pathway in Escherichia coli.
abstract: N-acetylneuraminic acid (NeuAc) has recently drawn much attention owing to its wide applications in many aspects. Besides extraction from natural materials, production of NeuAc was recently focused on enzymatic synthesis and whole-cell biocatalysis. In this study, we designed an artificial NeuAc biosynthetic pathway through intermediate N-acetylglucosamine 6-phosphate in Escherichia coli. In this pathway, N-acetylglucosamine 2-epimerase (slr1975) and glucosamine-6-phosphate acetyltransferase (GNA1) were heterologously introduced into E. coli from Synechocystis sp. PCC6803 and Saccharomyces cerevisiae EBY100, respectively. By derepressing the feedback inhibition of glucosamine-6-phosphate synthase, increasing the accumulation of N-acetylglucosamine and pyruvate, and blocking the catabolism of NeuAc, we were able to produce 1.62 g l⁻¹ NeuAc in recombinant E. coli directly from glucose. The NeuAc yield reached 7.85g l⁻¹ in fed-batch fermentation. This process offered an efficient fermentative method to produce NeuAc in microorganisms using glucose as carbon source and can be optimized for further improvement.
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, 27 Shanda Nanlu, Jinan 250100, PR China.
authors.forename: Junhua
authors.initials: J
authors.lastname: Kang
authors.forename: Pengfei
authors.initials: P
authors.lastname: Gu
authors.forename: Yang
authors.initials: Y
authors.lastname: Wang
authors.forename: Yikui
authors.initials: Y
authors.lastname: Li
authors.forename: Fan
authors.initials: F
authors.lastname: Yang
authors.forename: Qian
authors.initials: Q
authors.lastname: Wang
authors.forename: Qingsheng
authors.initials: Q
authors.lastname: Qi
mesh_terms: D000117:Acetylglucosamine / Q000031:analogs & derivatives* / Q000378:metabolism; D002238:Carbohydrate Epimerases / Q000235:genetics / Q000378:metabolism*; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000502:physiology*; D051081:Glucosamine 6-Phosphate N-Acetyltransferase / Q000235:genetics / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism*; D019158:N-Acetylneuraminic Acid / Q000096:biosynthesis* / Q000302:isolation & purification; D015202:Protein Engineering / Q000379:methods; D011994:Recombinant Proteins / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D046939:Synechocystis / Q000235:genetics* / Q000378:metabolism
chemical_list: D002352:Carrier Proteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C010004:N-acetylglucosamine 6-phosphate; C494969:GNA1 protein, S cerevisiae; D051081:Glucosamine 6-Phosphate N-Acetyltransferase; D002238:Carbohydrate Epimerases; C024519:N-acyl-D-glucosamine 2-epimerase; D019158:N-Acetylneuraminic Acid; D005947:Glucose; D000117:Acetylglucosamine