title: Improved production of trans-4-hydroxy-l-proline by chromosomal integration of the Vitreoscilla hemoglobin gene into recombinant Escherichia coli with expression of proline-4-hydroxylase.
abstract: trans-4-Hydroxy-l-proline (Hyp) is a chiral amino acid conventionally produced by acid hydrolysis of animal collagen, a process which involves the bottleneck problems of low efficiency and heavy environmental pollution. Biotransformation of l-proline into Hyp using recombinant whole-cell biocatalysis with proline-4-hydroxylase (P4H) is an environmentally-friendly alternative method. Since biohydroxylation of proline by whole cells is a high-oxygen-demand process, oxygen transfer needs to be improved. To solve this problem, the Vitreoscilla hemoglobin gene (vgb) was integrated into the chromosome of recombinant Escherichia coli expressing the P4H gene originally from Dactylosporangium sp. RH1. Expression of Vitreoscilla hemoglobin (VHb) resulted in a 94.4% increase of Hyp production in a 100-mL shaking flask culture compared to the same strain without VHb expression. Meanwhile in a fed-batch fermentation with a 1.4 L bioreactor, the expression of VHb led to an increase in Hyp production by 73.2% and biomass improved by 106%. We also found that acetic acid concentration was decreased by the expression of VHb during the fermentation. This work demonstrates that vgb chromosomal integration is an efficient way to improve Hyp production by enhancing oxygen transfer in recombinant E. coli.
authors: Key Laboratory for Industrial Biocatalysis, Ministry of Education of China, Institute of Biochemical Engineering, Department of Chemical Engineering, Tsinghua University, Beijing 100084, PR China; College of Chemistry and Life Science, Shenyang Normal University, Shenyang 110034, PR China.
authors: Tong-Xin
authors: TX
authors: Zhao
authors: Key Laboratory for Industrial Biocatalysis, Ministry of Education of China, Institute of Biochemical Engineering, Department of Chemical Engineering, Tsinghua University, Beijing 100084, PR China.
authors: Mei
authors: M
authors: Li
authors: Key Laboratory for Industrial Biocatalysis, Ministry of Education of China, Institute of Biochemical Engineering, Department of Chemical Engineering, Tsinghua University, Beijing 100084, PR China.
authors: Xiang
authors: X
authors: Zheng
authors: Key Laboratory for Industrial Biocatalysis, Ministry of Education of China, Institute of Biochemical Engineering, Department of Chemical Engineering, Tsinghua University, Beijing 100084, PR China.
authors: Cheng-Hua
authors: CH
authors: Wang
authors: Key Laboratory for Industrial Biocatalysis, Ministry of Education of China, Institute of Biochemical Engineering, Department of Chemical Engineering, Tsinghua University, Beijing 100084, PR China; Zhejiang Province Key Laboratory of Plant Secondary Metabolism and Regulation, College of Life Sciences, Zhejiang Sci-Tech University, Hangzhou 310018, PR China.
authors: Hong-Xin
authors: HX
authors: Zhao
authors: Key Laboratory for Industrial Biocatalysis, Ministry of Education of China, Institute of Biochemical Engineering, Department of Chemical Engineering, Tsinghua University, Beijing 100084, PR China. Electronic address: chongzhang@tsinghua.edu.cn.
authors: Chong
authors: C
authors: Zhang
authors: Key Laboratory for Industrial Biocatalysis, Ministry of Education of China, Institute of Biochemical Engineering, Department of Chemical Engineering, Tsinghua University, Beijing 100084, PR China.
authors: Xin-Hui
authors: XH
authors: Xing
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics*; D002876:Chromosomes, Bacterial / Q000235:genetics*; D004274:DNA, Recombinant / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D005285:Fermentation; D015870:Gene Expression; D005818:Genetic Engineering; D006909:Hydroxyproline / Q000096:biosynthesis*; D064828:Prolyl Hydroxylases / Q000235:genetics*; D054793:Truncated Hemoglobins / Q000235:genetics*; D020589:Vitreoscilla / Q000235:genetics*
chemical_list: D001426:Bacterial Proteins; D004274:DNA, Recombinant; D054793:Truncated Hemoglobins; C082613:hemoglobin protein, Vitreoscilla; D064828:Prolyl Hydroxylases; D006909:Hydroxyproline
keywords: Biocatalysis; Escherichia coli; Fermentation; Vitreoscilla hemoglobin; trans-4-Hydroxy-l-proline