a School of Engineering, Dalian Polytechnic University , Liaoning , Dalian , China.
b Marine Bioengineering Group, Dalian Institute of Chemical Physics , Chinese Academy of Sciences , Dalian , China.
Bioengineered. 2017 Nov 2;8(6):707-715. doi: 10.1080/21655979.2017.1317574. Epub 2017 May 19.
The production of poly-3-hydroxybutyrate (PHB) by photosynthetic cyanobacteria is a potentially sustainable production method for the biodegradable plastics industry. β-Ketoacyl-ACP reductase (FabG), from the photosynthetic cyanobacterium Synechocystis sp. PCC 6803 (SpFabG), is the first NADPH-dependent reductase in the fatty acid biosynthesis pathway. Its structure is similar to that of acetoacetyl-CoA reductase (SpPhaB), which is critical for PHB synthesis and can replace SpPhaB for acetoacetyl-CoA reduction in vitro. However, the specific function of SpFabG in fatty acid synthesis and whether SpFabG could participate in PHB synthesis in vivo were not yet clear. In this study, the role of SpFabG in fatty acid synthesis was first verified in vivo by knocking down and overexpressing of fabG. It was shown that SpFabG was essential yet not rate-limiting for fatty acid biosynthesis. The biochemical characterization of SpFabG using acetoacetyl-CoA as the substrate showed that the optimum temperature, optimum pH, K and k were 30°C, 7, 2.30 mM, and 19.85 s, respectively, which exemplified the ability of SpFabG to reduce acetoacetyl-CoA with a relatively low affinity and weak catalytic efficiency. Functional analysis of SpFabG in vivo indicated that SpFabG was able to partially complement SpPhaB under nitrogen-deprived conditions, and overexpression of fabG led to the diversion of partial carbon flux from fatty acid toward PHB synthesis.
聚 3-羟基丁酸酯 (PHB) 的光合蓝细菌生产是可生物降解塑料行业可持续生产的潜在方法。β-酮酰基-ACP 还原酶（FabG）来自光合蓝细菌集胞藻 6803（SpFabG），是脂肪酸生物合成途径中的第一个 NADPH 依赖性还原酶。其结构类似于乙酰乙酰辅酶 A 还原酶（SpPhaB），这对于 PHB 合成至关重要，并且可以在体外替代 SpPhaB 还原乙酰乙酰辅酶 A。然而，SpFabG 在脂肪酸合成中的具体功能以及 SpFabG 是否可以在体内参与 PHB 合成尚不清楚。在本研究中，首先通过敲低和过表达 fabG 在体内验证了 SpFabG 在脂肪酸合成中的作用。结果表明，SpFabG 对于脂肪酸生物合成是必需的，但不是限速步骤。使用乙酰乙酰辅酶 A 作为底物对 SpFabG 的生化特性进行了表征，结果表明最适温度、最适 pH 值、K 和 k 分别为 30°C、7、2.30 mM 和 19.85 s，这说明了 SpFabG 以相对较低的亲和力和较弱的催化效率还原乙酰乙酰辅酶 A 的能力。SpFabG 在体内的功能分析表明，SpFabG 能够在氮饥饿条件下部分补充 SpPhaB，而过表达 fabG 导致部分碳通量从脂肪酸转向 PHB 合成。
