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Engineering of Bacterial Methyl Ketone Synthesis for Biofuels

机译：用于生物燃料的细菌甲基酮合成工程

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We have engineered Escherichia coli to overproduce saturated and monounsaturated aliphatic methyl ketones in the C_(11) to C_(15) (diesel) range; this group of methyl ketones includes 2-undecanone and 2-tridecanone, which are of importance to the flavor and fragrance industry and also have favorable cetane numbers (as we report here). We describe specific improvements that resulted in a 700-fold enhancement in methyl ketone titer relative to that of a fatty acid-overproducing E. coli strain, including the following: (i) overproduction of β-ketoacyl coenzyme A (CoA) thioesters achieved by modification of the β-oxidation pathway (specifically, overexpression of a heterologous acyl-CoA oxidase and native FadB and chromosomal deletion of fadA ) and (ii) overexpression of a native thioesterase (FadM). FadM was previously associated with oleic acid degradation, not methyl ketone synthesis, but outperformed a recently identified methyl ketone synthase ( Solanum habrochaites MKS2 [ Sh MKS2], a thioesterase from wild tomato) in β-ketoacyl-CoA-overproducing strains tested. Whole-genome transcriptional (microarray) studies led to the discovery that FadM is a valuable catalyst for enhancing methyl ketone production. The use of a two-phase system with decane enhanced methyl ketone production by 4- to 7-fold in addition to increases from genetic modifications.
机译：我们对大肠杆菌进行了改造，使其过量生产C_（11）至C_（15）（柴油）范围内的饱和和单不饱和脂肪族甲基酮;这组甲基酮包括2-十一碳烷酮和2-tricancanone，它们对香料和香料行业很重要，而且十六烷值也很有利（如我们在此报告）。我们描述了导致甲基酮滴度相对于脂肪酸过量生产的大肠杆菌菌株增加700倍的特定改进，包括以下方面：（i）通过以下方法实现的β-酮酰基辅酶A（CoA）硫酯的过量生产β氧化途径的修饰（特别是异源酰基辅酶A氧化酶和天然FadB的过表达以及fadA的染色体缺失）和（ii）天然硫酯酶（FadM）的过表达。 FadM以前与油酸降解有关，而不与甲基酮合成有关，但在测试过的β-酮酰基-CoA高产菌株中，FadM的表现优于最近鉴定出的甲基酮合酶（茄子茄子MKS2 [Sh MKS2]，一种来自野生番茄的硫酯酶）。全基因组转录（微阵列）研究导致发现FadM是增强甲基酮生产的有价值的催化剂。与癸烷一起使用的两相系统，除了遗传修饰的增加之外，还将甲基酮的产量提高了4到7倍。

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