title: Enzymatic process optimization for the in vitro production of isoprene from mevalonate.
abstract: BACKGROUND
  As an important bulk chemical for synthetic rubber, isoprene can be biosynthesized by robust microbes. But rational engineering and optimization are often demanded to make the in vivo process feasible due to the complexities of cellular metabolism. Alternative synthetic biochemistry strategies are in fast development to produce isoprene or isoprenoids in vitro.
  
  
  RESULTS
  This study set up an in vitro enzyme synthetic chemistry process using 5 enzymes in the lower mevalonate pathway to produce isoprene from mevalonate. We found the level and ratio of individual enzymes would significantly affect the efficiency of the whole system. The optimized process using 10 balanced enzyme unites (5.0 µM of MVK, PMK, MVD; 10.0 µM of IDI, 80.0 µM of ISPS) could produce 6323.5 µmol/L/h (430 mg/L/h) isoprene in a 2 ml in vitro system. In a scale up process (50 ml) only using 1 balanced enzyme unit (0.5 µM of MVK, PMK, MVD; 1.0 µM of IDI, 8.0 µM of ISPS), the system could produce 302 mg/L isoprene in 40 h, which showed higher production rate and longer reaction phase with comparison of the in vivo control.
  
  
  CONCLUSIONS
  By optimizing the enzyme levels of lower MVA pathway, synthetic biochemistry methods could be set up for the enzymatic production of isoprene or isoprenoids from mevalonate.
authors.affiliation: CAS Key Laboratory of Bio-based Materials, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, No. 189 Songling Road, Laoshan District, Qingdao, 266101, China.|State Key Laboratory Base of Eco-Chemical Engineering, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao, 266042, China.
authors.forename: Tao
authors.initials: T
authors.lastname: Cheng
authors.affiliation: CAS Key Laboratory of Bio-based Materials, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, No. 189 Songling Road, Laoshan District, Qingdao, 266101, China.
authors.forename: Hui
authors.initials: H
authors.lastname: Liu
authors.affiliation: CAS Key Laboratory of Bio-based Materials, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, No. 189 Songling Road, Laoshan District, Qingdao, 266101, China. huibinzou@hotmail.com.|College of Chemical Engineering, Qingdao University of Science and Technology, Qingdao, 266042, China. huibinzou@hotmail.com.
authors.forename: Huibin
authors.initials: H
authors.lastname: Zou
authors.affiliation: College of Chemical Engineering, Qingdao University of Science and Technology, Qingdao, 266042, China.
authors.forename: Ningning
authors.initials: N
authors.lastname: Chen
authors.affiliation: College of Chemical Engineering, Qingdao University of Science and Technology, Qingdao, 266042, China.
authors.forename: Mengxun
authors.initials: M
authors.lastname: Shi
authors.affiliation: State Key Laboratory Base of Eco-Chemical Engineering, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao, 266042, China.
authors.forename: Congxia
authors.initials: C
authors.lastname: Xie
authors.affiliation: CAS Key Laboratory of Bio-based Materials, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, No. 189 Songling Road, Laoshan District, Qingdao, 266101, China. zhaoguang@qibebt.ac.cn.
authors.forename: Guang
authors.initials: G
authors.lastname: Zhao
authors.affiliation: CAS Key Laboratory of Bio-based Materials, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, No. 189 Songling Road, Laoshan District, Qingdao, 266101, China. xianmo@qibebt.ac.cn.
authors.forename: Mo
authors.initials: M
authors.lastname: Xian
mesh_terms: D002070:Butadienes; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D045782:Hemiterpenes / Q000096:biosynthesis*; D060847:Metabolic Engineering / Q000379:methods*; D008798:Mevalonic Acid / Q000378:metabolism*; D010420:Pentanes; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism
chemical_list: D002070:Butadienes; D045782:Hemiterpenes; D010420:Pentanes; C005059:isoprene; D008798:Mevalonic Acid
keywords: Bio-based chemicals; Enzymatic process; Isoprene; Isoprenoids; Mevalonate; Synthetic biochemistry