title: Combinatorial analysis of enzymatic bottlenecks of L-tyrosine pathway by p-coumaric acid production in Saccharomyces cerevisiae.
abstract: OBJECTIVE
  To identify new enzymatic bottlenecks of L-tyrosine pathway for further improving the production of L-tyrosine and its derivatives.
  
  
  RESULTS
  When ARO4 and ARO7 were deregulated by their feedback resistant derivatives in the host strains, the ARO2 and TYR1 genes, coding for chorismate synthase and prephenate dehydrogenase were further identified as new important rate-limiting steps. The yield of p-coumaric acid in the feedback-resistant strain overexpressing ARO2 or TYR1, was significantly increased from 6.4 to 16.2 and 15.3 mg l-1, respectively. Subsequently, we improved the strain by combinatorial engineering of pathway genes increasing the yield of p-coumaric acid by 12.5-fold (from 1.7 to 21.3 mg l-1) compared with the wild-type strain. Batch cultivations revealed that p-coumaric acid production was correlated with cell growth, and the formation of by-product acetate of the best producer NK-M6 increased to 31.1 mM whereas only 19.1 mM acetate was accumulated by the wild-type strain.
  
  
  CONCLUSIONS
  Combinatorial metabolic engineering provides a new strategy for further improvement of L-tyrosine or other metabolic biosynthesis pathways in S. cerevisiae.
authors.affiliation: The Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Nankai University, No. 94 Weijin Road, Nankai Distract, Tianjin, 300071, People's Republic of China.
authors.forename: Jiwei
authors.initials: J
authors.lastname: Mao
authors.affiliation: The Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Nankai University, No. 94 Weijin Road, Nankai Distract, Tianjin, 300071, People's Republic of China.
authors.forename: Quanli
authors.initials: Q
authors.lastname: Liu
authors.affiliation: The Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Nankai University, No. 94 Weijin Road, Nankai Distract, Tianjin, 300071, People's Republic of China.
authors.forename: Xiaofei
authors.initials: X
authors.lastname: Song
authors.affiliation: The Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Nankai University, No. 94 Weijin Road, Nankai Distract, Tianjin, 300071, People's Republic of China.
authors.forename: Hesuiyuan
authors.initials: H
authors.lastname: Wang
authors.affiliation: Environmental Protection Technical Development Center, Tianjin, 300191, People's Republic of China.
authors.forename: Hui
authors.initials: H
authors.lastname: Feng
authors.affiliation: The Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Nankai University, No. 94 Weijin Road, Nankai Distract, Tianjin, 300071, People's Republic of China.
authors.forename: Haijin
authors.initials: H
authors.lastname: Xu
authors.affiliation: The Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Nankai University, No. 94 Weijin Road, Nankai Distract, Tianjin, 300071, People's Republic of China. mingqiangqiao@aliyun.com.
authors.forename: Mingqiang
authors.initials: M
authors.lastname: Qiao
mesh_terms: D000890:Anti-Infective Agents / Q000378:metabolism*; D003373:Coumaric Acids; D015870:Gene Expression; D060847:Metabolic Engineering; D053858:Metabolic Networks and Pathways / Q000235:genetics; D019761:Phosphorus-Oxygen Lyases / Q000235:genetics / Q000378:metabolism*; D011303:Prephenate Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D011422:Propionates / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism*
chemical_list: D000890:Anti-Infective Agents; D003373:Coumaric Acids; D011422:Propionates; D029701:Saccharomyces cerevisiae Proteins; D014443:Tyrosine; D011303:Prephenate Dehydrogenase; C019895:chorismate synthase; D019761:Phosphorus-Oxygen Lyases; C495469:p-coumaric acid
keywords: ARO2; Gene overexpression; L-Tyrosine; Saccharomyces cerevisiae; TYR1; p-Coumaric acid