Roles of the FabA and FabZ beta-hydroxyacyl-acyl carrier protein dehydratases in Escherichia coli fatty acid biosynthesis
Roles of the FabA and FabZ beta-hydroxyacyl-acyl carrier protein dehydratases in Escherichia coli fatty acid biosynthesis
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DOI：
10.1074/jbc.271.44.27795
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发表时间：
1996-11-01
影响因子：
4.8
通讯作者：
Rock, CO
中科院分区：
文献类型：
--
作者：
Heath, RJ;Rock, CO
有两个基因，fabA和fabZ，编码β-羟酰基-酰基载体蛋白（ACP）脂肪酶，在大肠杆菌的解离的II型脂肪酸合酶系统中发挥作用。我们已经研究了它们在脂肪酸合成中的作用，通过纯化这两种蛋白质，并使用其他五种纯化的蛋白质在体外重建脂肪酸合成循环。FabA和FabZ表现出广泛的、重叠的链长特异性，FabZ淀粉酶有效地催化短链β-羟酰基-ACP和长链饱和和不饱和β-羟酰基-ACP的脱水。FabA对中等链长的β-羟酰基-ACP最具活性，并且对短链和长链饱和的β-羟酰基-ACP也具有显著的活性。值得注意的是，FabA在长链不饱和β-羟酰基-ACP的脱水中几乎是无活性的。仅在β-酮脂酰-ACP合酶I（FabB）存在下检测到FabA在脂肪酸合成的10-碳阶段引入双键。酵母双杂交分析未能检测到FabA和FabB之间的相互作用，因此FabB向不饱和脂肪酸合成的中间体的通道归因于缩合酶对顺式癸烯酰基-ACP的亲和力。FabZ的广泛底物特异性与FabA对长链不饱和β-羟酰基-ACP的无活性相结合，为具有遗传改变的两种β-羟酰基转移酶水平的细胞的表型提供了生物化学解释。
There are two genes, fabA and fabZ, encoding beta-hydroxyacyl-acyl carrier protein (ACP) dehydratases that function in the dissociated, type II fatty acid synthase system of Escherichia coli. We have investigated their roles in fatty acid synthesis by purifying the two proteins and reconstituting cycles of fatty acid synthesis in vitro using five other purified proteins. FabA and FabZ exhibited broad, overlapping chain length specificities, The FabZ dehydratase efficiently catalyzed the dehydration of short chain beta-hydroxyacyl-ACPs and long chain saturated and unsaturated beta-hydroxyacyl-ACPs. FabA was most active on intermediate chain length beta-hydroxyacyl-ACPs and also possessed significant activity toward both short and long chain saturated beta-hydroxyacyl-ACPs. Significantly, FabA was virtually inactive in the dehydration of long chain unsaturated beta-hydroxyacyl-ACP. The introduction of the double bond at the 10-carbon stage of fatty acid synthesis by FabA was only detected in the presence of beta-ketoacyl-ACP synthase I (FabB). A yeast two-hybrid analysis failed to detect an interaction between FabA and FabB, therefore the channeling of intermediates toward unsaturated fatty acid synthesis by FabB was attributed to the affinity of the condensing enzyme for cis-decenoyl-ACP. The broad substrate specificity of FabZ coupled with the inactivity of FabA toward a long chain unsaturated p hydroxyacyl-ACP provides a biochemical explanation for the phenotypes of cells with genetically altered levels of the two dehydratases.