title: Improvement of amorpha-4,11-diene production by a yeast-conform variant of Vitreoscilla hemoglobin.
abstract: Amorpha-4,11-diene is the precursor of the antimalarial compound artemisinin. The effect of Vitreoscilla hemoglobin (VHb) and its yeast-conform variant (VHbm) on amorpha-4,11-diene production in engineered Saccharomyces cerevisiae was investigated. First, the VHb gene was mutated to the yeast-conform variant VHbm based on step-by-step extension of a short region of the gene through a series of polymerase chain reactions (PCR). The artificial VHbm gene contained codons preferred by the yeast translation machinery. Two yeast expression vectors containing VHb or VHbm gene were constructed and introduced into the amorpha-4,11-diene-producing strain S. cerevisiae WK1 to form WK1[VHb] and WK1[VHbm], respectively. Western blot and CO-difference spectrum absorbance assay showed that VHb and VHbm were successfully expressed. In shake flasks, VHbm expression conferred higher cell growth than VHb expression. GC-MS results indicated the amorpha-4,11-diene production in WK1[VHbm] and WK1[VHb] was 3- and 2-fold higher than that in WK1, respectively. This suggests that VHb might improve the amorpha-4,11-diene production in engineered S. cerevisiae.
authors.affiliation: Institute of Materia Medica, Chinese Academy of Medical Sciences & Peking Union Medical College (State Key Laboratory of Bioactive Substance and Function of Natural Medicines & Ministry of Health Key Laboratory of Biosynthesis of Natural Products), Beijing 100050, China.
authors.forename: Junhao
authors.initials: J
authors.lastname: Shen
authors.forename: Haina
authors.initials: H
authors.lastname: Zheng
authors.forename: Xiaohui
authors.initials: X
authors.lastname: Zhi
authors.forename: Yanwei
authors.initials: Y
authors.lastname: Shi
authors.forename: Yong
authors.initials: Y
authors.lastname: Huang
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.forename: Yongqin
authors.initials: Y
authors.lastname: Chen
authors.forename: Jianqiang
authors.initials: J
authors.lastname: Kong
authors.forename: Ping
authors.initials: P
authors.lastname: Zhu
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D015153:Blotting, Western; D017931:DNA Primers; D008969:Molecular Sequence Data; D010957:Plasmids; D000081223:Polycyclic Sesquiterpenes; D016133:Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid; D012717:Sesquiterpenes / Q000378:metabolism*; D054793:Truncated Hemoglobins / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D017931:DNA Primers; D000081223:Polycyclic Sesquiterpenes; D012717:Sesquiterpenes; D054793:Truncated Hemoglobins; C515348:amorpha-4,11-diene; C082613:hemoglobin protein, Vitreoscilla