Overexpression of endogenous 1-deoxy-d-xylulose 5-phosphate synthase (DXS) in cyanobacterium Synechocystis sp. PCC6803 accelerates protein aggregation

• Published in: Journal of bioscience and bioengineering 2017-05, Vol.123 (5), p.590-596
• Main Authors: Kudoh, Kai, Hotta, Shingo, Sekine, Midori, Fujii, Rintaro, Uchida, Arisu, Kubota, Genma, Kawano, Yusuke, Ihara, Masaki
• Format: Article
• Language: English
• Subjects: 1-Deoxyxylulose 5-phosphate synthase; Alkyl and Aryl Transferases - biosynthesis; Alkyl and Aryl Transferases - genetics; Alkyl and Aryl Transferases - metabolism; Butadienes; Erythritol - analogs & derivatives; Erythritol - metabolism; Gases - metabolism; Hemiterpenes - biosynthesis; Isoprene; MEP pathway; Metabolic Engineering; Pentanes; Pentosephosphates - biosynthesis; Protein Aggregates; Protein solubility; RNA, Messenger - analysis; Solubility; Sugar Phosphates - metabolism; Synechocystis - genetics; Synechocystis - metabolism; Terpenes - metabolism; Transferases - biosynthesis; Transferases - genetics; Transferases - metabolism
• ISSN: 1389-1723; 1347-4421
• DOI: 10.1016/j.jbiosc.2017.01.001

1-Deoxy-d-xylulose 5-phosphate synthase (DXS) is a rate-limiting enzyme in the 2-C-methyl-d-erythritol 4-phosphate (MEP) pathway, which is responsible for the production of precursors of all isoprenoids. In a previous study, we had examined the overexpression of an endogenous DXS in a Synechocystis sp. PCC6803 mutant (DXS_ox), and found that the dxs mRNA level was 4-fold higher than that in the wild-type (WT) strain. However, the DXS protein level was only 1.5-fold higher, leading to the assumption that the level might be regulated by post-transcriptional events. In this study, we have additionally introduced an exogenous isoprene synthase (IspS; which can release MEP pathway products from the cell as gaseous isoprene) into the WT and DXS_ox strains (WT-isP and DXSox-isP strains, respectively), and their detailed DXS expression profiles were investigated from the induction phase through to the late-logarithmic phase. In the induction phase, the isoprene productivity of the DXSox-isP strain was slightly but significantly (1.4- to 1.8-fold) higher than that of the WT-isP strain, whereas the levels were comparable in the other phases. Interestingly, the ratios of soluble:insoluble DXS protein were remarkably low in the DXSox-isP strain during the induction phase to the early-logarithmic phase, resulting in a moderate level of soluble DXS. All our results suggested that the high translation rate of DXS disturbs the refolding process of DXS. To enhance the concentration of the active DXS in cyanobacteria, the enhancement of the DXS maturation system or the introduction of exogenous and robust DXS proteins might be necessary.