PDF(Pubmed)
Abstract：
Sedoheptulose 7-phosphate (SH7P) cyclases are a subset of sugar phosphate cyclases that are known to catalyze the first committed step in many biosynthetic pathways in primary and secondary metabolism. Among them are 2-epi-5-epi-valiolone synthase (EEVS) and 2-epi-valiolone synthase (EVS), two closely related SH7P cyclases that catalyze the conversion of SH7P to 2-epi-5-epi-valiolone and 2-epi-valiolone, respectively. However, how these two homologous enzymes use a common substrate to produce stereochemically different products is unknown. Two competing hypotheses have been proposed for the stereospecificity of EEVS and EVS: (1) variation in aldol acceptor geometry during enzyme catalysis, and (2) preselection of the α-pyranose or β-pyranose forms of the substrate by the enzymes. Yet, there is no direct evidence to support or rule out either of these hypotheses. Here we report the synthesis of the carba-analogs of the α-pyranose and β-pyranose forms of SH7P and their use in probing the stereospecificity of ValA (EEVS from Streptomyces hygroscopicus subsp. jinggangensis) and Amir_2000 (EVS from Actinosynnema mirum DSM 43827). Kinetic studies of the enzymes in the presence of the synthetic compounds as well as docking studies of the enzymes with the α- and β-pyranose forms of SH7P suggest that the inverted configuration of the products of EEVS and EVS is not due to the preselection of the different forms of the substrate by the enzymes.
摘要：
Sedo庚酮糖7-磷酸(SH7P)环化酶是已知催化初级和次级代谢中许多生物合成途径的第一个关键步骤的糖磷酸盐环化酶的子集。其中包括2-epi-5-epi-Valiolone合酶（EEVS）和2-epi-Valiolone合酶（EVS），两种密切相关的SH7P环化酶，催化SH7P转化为2-epi-5-epi-Valiolone和2-epi-Valiolone，分别。然而,这两种同源酶如何使用一个共同的底物来产生不同的立体化学产物是未知的。针对EEVS和EVS的立体特异性提出了两个竞争性假设：（1）酶催化过程中醛醇受体几何形状的变化，和(2)通过酶预选α-吡喃糖或β-吡喃糖形式的底物。然而，没有直接证据支持或排除这些假设。在这里，我们报告了SH7P的α-吡喃糖和β-吡喃糖形式的碳类似物的合成及其在探索ValA立体特异性中的用途（来自吸湿性链霉菌亚种的EEVS。井冈）和Amir_2000（来自mirum放线菌DSM43827的EVS）。在合成化合物存在下的酶的动力学研究以及酶与SH7P的α-和β-吡喃糖形式的对接研究表明，EEVS和EVS产物的反向构型不是由于酶对不同形式的底物的预选。
