title: Dynamic control of the mevalonate pathway expression for improved zeaxanthin production in Escherichia coli and comparative proteome analysis.
abstract: Engineered heterologous multi-gene metabolic pathways often suffer from flux imbalance and toxic metabolites, as the production host typically lacks the regulatory mechanisms for the heterologous pathway. Here, we first coordinated the expression of all genes of the mevalonate (MEV) pathway from Saccharomyces cerevisiae using the tunable intergenic regions (TIGRs), and then dynamically regulated the TIGR-mediated MEV pathway to prevent the accumulation of toxic metabolites by using IPP/FPP-responsive promoter. After introduction of the dynamically controlled TIGR-mediated MEV pathway into Escherichia coli, the content and concentration of zeaxanthin in shaker flask cultures were 2.0- and 2.1-fold higher, respectively, than those of the strain harboring the statically controlled non-TIGR-mediated MEV pathway. The content and concentration of zeaxanthin in E. coli ZEAX (pZSPgadE-MevTTIGR-MevBTIGRIS-2) reached 722.46mg/L and 23.16mg/g dry cell weight (DCW), respectively, in 5.0L fed-batch fermentation. We also comparatively analyzed the proteomes between E. coli ZEAX and E. coli ZEAX (pZSPgadE-MevTTIGR-MevBTIGRIS-2) to understand the mechanism of zeaxanthin biosynthesis. The results of the comparative proteomes demonstrate that zeaxanthin overproduction may be associated with increased precursor availability, increased NADPH availability, increased ATP availability, oxidative stress response, and increased membrane storage capacity for zeaxanthin due to changes in both cellular shape and membrane composition.
authors.affiliation: Biotechnology Research Center and Biomedical Center, South China Sea Bio-Resource Exploitation and Utilization Collaborative Innovation Center, School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China.
authors.forename: Hong-Jie
authors.initials: HJ
authors.lastname: Shen
authors.affiliation: Biotechnology Research Center and Biomedical Center, South China Sea Bio-Resource Exploitation and Utilization Collaborative Innovation Center, School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China.
authors.forename: Bi-Yan
authors.initials: BY
authors.lastname: Cheng
authors.affiliation: Biotechnology Research Center and Biomedical Center, South China Sea Bio-Resource Exploitation and Utilization Collaborative Innovation Center, School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China.
authors.forename: Yan-Mei
authors.initials: YM
authors.lastname: Zhang
authors.affiliation: Biotechnology Research Center and Biomedical Center, South China Sea Bio-Resource Exploitation and Utilization Collaborative Innovation Center, School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China.
authors.forename: Liang
authors.initials: L
authors.lastname: Tang
authors.affiliation: Biotechnology Research Center and Biomedical Center, South China Sea Bio-Resource Exploitation and Utilization Collaborative Innovation Center, School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China.
authors.forename: Zhen
authors.initials: Z
authors.lastname: Li
authors.affiliation: Biotechnology Research Center and Biomedical Center, South China Sea Bio-Resource Exploitation and Utilization Collaborative Innovation Center, School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China.
authors.forename: Yi-Fan
authors.initials: YF
authors.lastname: Bu
authors.affiliation: Biotechnology Research Center and Biomedical Center, South China Sea Bio-Resource Exploitation and Utilization Collaborative Innovation Center, School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China.
authors.forename: Xi-Ran
authors.initials: XR
authors.lastname: Li
authors.affiliation: Biotechnology Research Center and Biomedical Center, South China Sea Bio-Resource Exploitation and Utilization Collaborative Innovation Center, School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China.
authors.forename: Gui-Qiao
authors.initials: GQ
authors.lastname: Tian
authors.affiliation: Biotechnology Research Center and Biomedical Center, South China Sea Bio-Resource Exploitation and Utilization Collaborative Innovation Center, School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China. Electronic address: lssljz@mail.sysu.edu.cn.
authors.forename: Jian-Zhong
authors.initials: JZ
authors.lastname: Liu
mesh_terms: D004926:Escherichia coli / Q000502:physiology*; D005285:Fermentation / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics; D060847:Metabolic Engineering / Q000379:methods*; D064688:Metabolic Flux Analysis / Q000379:methods*; D053858:Metabolic Networks and Pathways / Q000502:physiology; D008798:Mevalonic Acid / Q000378:metabolism*; D008954:Models, Biological; D011401:Promoter Regions, Genetic / Q000235:genetics; D020543:Proteome / Q000235:genetics / Q000378:metabolism*; D065146:Zeaxanthins / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D020543:Proteome; D065146:Zeaxanthins; D008798:Mevalonic Acid
keywords: Dynamic regulation; Escherichia coli; Proteome; TIGR; Zeaxanthin; Zeaxanthin (PubChem CID: 5280899)