title: Heterologous pathway for the production of L-phenylglycine from glucose by E. coli.
abstract: The aproteinogenic amino acid L-phenylglycine (L-Phg) is an important side chain building block for the preparation of several antibiotics and taxol. To biosynthesis L-Phg from glucose, an engineered Escherichia coli containing L-Phg synthetic genes was firstly developed by an L-phenylalanine producing chassis supplying phenylpyruvate. The enzymes HmaS (L-4-hydroxymandelate synthase), Hmo (L-4-hydroxymandelate oxidase) and HpgT (L-4-hydroxyphenylglycine transaminase) from Amycolatopsis orientalis as well as Streptomyces coelicolor were heterologously expressed in E. coli and purified to evaluate their abilities on L-Phg formation. HpgT conversing phenylglyoxylate to L-Phg uses an unusual amino donor L-phenylalanine, which releases another phenylpyruvate as the substrate of HmaS. Thus, a recycle reaction was developed to maximize the utilization of precursor phenylpyruvate. To amplify the accumulation of L-Phg, the effects of attenuating L-phenylalanine transamination was investigated. After deletion of tyrB and aspC, L-Phg yield increased by 12.6-fold. The limiting step in the L-Phg biosynthesis was also studied; the L-Phg yield was further improved by 14.9-fold after enhancing hmaS expression. Finally, by optimizing expression of hmaS, hmo and hpgT and attenuation of L-phenylalanine transamination, the L-Phg yield was increased by 224-fold comparing with the original strain.
authors.affiliation: The Key Laboratory of Industrial Biotechnology, Ministry of Education, National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China. Electronic address: liushuangping668@126.com.
authors.forename: Shuang Ping
authors.initials: SP
authors.lastname: Liu
authors.affiliation: The Key Laboratory of Industrial Biotechnology, Ministry of Education, National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China.
authors.forename: Rui Xia
authors.initials: RX
authors.lastname: Liu
authors.affiliation: The Key Laboratory of Industrial Biotechnology, Ministry of Education, National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China.
authors.forename: Ashraf A M M
authors.initials: AA
authors.lastname: El-Rotail
authors.affiliation: The Key Laboratory of Industrial Biotechnology, Ministry of Education, National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China.
authors.forename: Zhong Yang
authors.initials: ZY
authors.lastname: Ding
authors.affiliation: The Key Laboratory of Industrial Biotechnology, Ministry of Education, National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China.
authors.forename: Zheng Hua
authors.initials: ZH
authors.lastname: Gu
authors.affiliation: The Key Laboratory of Industrial Biotechnology, Ministry of Education, National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China. Electronic address: zhangl@jiangnan.edu.cn.
authors.forename: Liang
authors.initials: L
authors.lastname: Zhang
authors.affiliation: The Key Laboratory of Industrial Biotechnology, Ministry of Education, National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China. Electronic address: gyshi@jiangnan.edu.cn.
authors.forename: Gui Yang
authors.initials: GY
authors.lastname: Shi
mesh_terms: D000192:Actinomycetales / Q000201:enzymology / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D005998:Glycine / Q000031:analogs & derivatives* / Q000032:analysis / Q000378:metabolism; D053858:Metabolic Networks and Pathways; D010649:Phenylalanine / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D013302:Streptomyces / Q000201:enzymology / Q000235:genetics; D000637:Transaminases / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; D010649:Phenylalanine; C008852:2-phenylglycine; D000637:Transaminases; D005947:Glucose; D005998:Glycine
keywords: Amycolatopsis orientalis; Streptomyces coelicolor; l-4-Hydroxyphenylglycine aminotransferase; l-Phenylglycine