<br>大肠杆菌不饱和脂肪酸合成：fabA 基因的复杂转录以及 FabB 催化的基本反应的体内鉴定。,Journal of Biological Chemistry

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大肠杆菌不饱和脂肪酸合成：fabA 基因的复杂转录以及 FabB 催化的基本反应的体内鉴定。
Journal of Biological Chemistry ( IF 4.0 ) Pub Date : 2009 Oct 23 , DOI: 10.1074/jbc.m109.023440
Youjun Feng ₁ , John E Cronan

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尽管大肠杆菌的不饱和脂肪酸（UFA）合成途径是此类途径的原型，但多年来积累了一些未解决的问题。关键的参与者是 fabA 和 fabB 基因。早期对 fabA 转录的研究表明，该基因是从两个启动子转录的，其中一个启动子受到 FadR 蛋白的正向调控。另一个较弱的启动子（无法用当时可用的技术进行定位）被认为是组成型的，因为它的功能独立于 FadR。然而，最近发现 FabR 负调节因子可以抑制 fabA 转录。我们报告弱启动子与 FadR 依赖性启动子重叠并受 FabR 调节。该启动子在迄今为止测序的所有大肠杆菌和沙门氏菌基因组中都是严格保守的，并且被认为可以防止外源饱和脂肪酸对 fabA 转录的不当调节。此外，fabAup 启动子（之前通过选择增加 FabA 活性而分离的突变启动子）被证明是通过位于 fabA 紧邻上游的基因内的四个碱基缺失从头创建的启动子。尽管已知 FabB 可以催化延伸反应，但对 FabB 催化的关键 UFA 合成反应的论证一直难以捉摸。缺乏 FabB 的菌株是 UFA 营养缺陷型，表明该酶催化 UFA 合成中的一个重要步骤。使用特异性水解短链酰基-ACP 的硫酯酶，首次在体内追踪 UFA 合成途径的中间体。这些实验表明，fabB突变菌株比亲本野生型菌株积累了更少的顺-5-十二碳烯酸。这些数据表明，FabB 催化的 UFA 合成中的关键反应是 FabA 产生的 cis-3-decenoyl-ACP 的延伸。

"点击查看英文标题和摘要"

Escherichia coli unsaturated fatty acid synthesis: complex transcription of the fabA gene and in vivo identification of the essential reaction catalyzed by FabB.

Although the unsaturated fatty acid (UFA) synthetic pathway of Escherichia coli is the prototype of such pathways, several unresolved issues have accumulated over the years. The key players are the fabA and fabB genes. Earlier studies of fabA transcription showed that the gene was transcribed from two promoters, with one being positively regulated by the FadR protein. The other weaker promoter (which could not be mapped with the technology then available) was considered constitutive because its function was independent of FadR. However, the FabR negative regulator was recently shown to represses fabA transcription. We report that the weak promoter overlaps the FadR-dependent promoter and is regulated by FabR. This promoter is strictly conserved in all E. coli and Salmonella enterica genomes sequenced to date and is thought to provide insurance against inappropriate regulation of fabA transcription by exogenous saturated fatty acids. Also, the fabAup promoter, a mutant promoter previously isolated by selection for increased FabA activity, was shown to be a promoter created de novo by a four-base deletion within the gene located immediately upstream of fabA. Demonstration of the key UFA synthetic reaction catalyzed by FabB has been elusive, although it was known to catalyze an elongation reaction. Strains lacking FabB are UFA auxotrophs indicating that the enzyme catalyzes an essential step in UFA synthesis. Using thioesterases specific for hydrolysis of short chain acyl-ACPs, the intermediates of the UFA synthetic pathway have been followed in vivo for the first time. These experiments showed that a fabB mutant strain accumulated less cis-5-dodecenoic acid than the parental wild-type strain. These data indicate that the key reaction in UFA synthesis catalyzed by FabB is elongation of the cis-3-decenoyl-ACP produced by FabA.

更新日期：2017-01-31

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