Departamento de Biología Funcional e Instituto Universitario de Oncología del Principado de Asturias (I.U.O.P.A), Universidad de Oviedo, Oviedo, Spain.
Metab Eng. 2013 Nov;20:187-97. doi: 10.1016/j.ymben.2013.10.002. Epub 2013 Oct 19.
Mithramycin (MTM) is a polyketide antitumor compound produced by Streptomyces argillaceus constituted by a tricyclic aglycone with two aliphatic side chains, a trisaccharide and a disaccharide chain. The biosynthesis of the polyketide aglycone is initiated by the condensation of ten malonyl-CoA units to render a carbon chain that is modified to a tetracyclic intermediate and sequentially glycosylated by five deoxysugars originated from glucose-1-phosphate. Further oxidation and reduction render the final compound. We aimed to increase the precursor supply of malonyl-CoA and/or glucose-1-phosphate in S. argillaceus to enhance MTM production. We have shown that by overexpressing either the S. coelicolor phosphoglucomutase gene pgm or the acetyl-CoA carboxylase ovmGIH genes from the oviedomycin biosynthesis gene cluster in S. argillaceus, we were able to increase the intracellular pool of glucose-1-phosphate and malonyl-CoA, respectively. Moreover, we have cloned the S. argillaceus ADP-glucose pyrophosphorylase gene glgCa and the acyl-CoA:diacylglycerol acyltransferase gene aftAa, and we showed that by inactivating them, an increase of the intracellular concentration of glucose-1-phosphate/glucose-6-phosphate and malonyl-CoA/acetyl-CoA was observed, respectively. Each individual modification resulted in an enhancement of MTM production but the highest production level was obtained by combining all strategies together. In addition, some of these strategies were successfully applied to increase production of four MTM derivatives with improved pharmacological properties: demycarosyl-mithramycin, demycarosyl-3D-β-D-digitoxosyl-mithramycin, mithramycin SK and mithramycin SDK.
密特霉素（MTM）是一种聚酮类抗肿瘤化合物，由链霉菌（Streptomyces argillaceus）产生，由具有两个脂族侧链的三环糖苷组成，一个三糖和一个二糖链。该聚酮类糖苷的生物合成由十个丙二酰辅酶 A 单元的缩合起始，生成一条经修饰的四环中间体，并由五个源自葡萄糖-1-磷酸的脱氧糖依次糖基化。进一步的氧化和还原生成最终化合物。我们旨在增加链霉菌中丙二酰辅酶 A 和/或葡萄糖-1-磷酸的前体供应，以提高 MTM 的产量。我们已经表明，通过在链霉菌中过表达来自奥维霉素生物合成基因簇的链霉菌磷酸葡萄糖变位酶基因 pgm 或乙酰辅酶 A 羧化酶 ovmGIH 基因，我们能够分别增加细胞内葡萄糖-1-磷酸和丙二酰辅酶 A 的池。此外，我们克隆了链霉菌 ADP-葡萄糖焦磷酸化酶基因 glgCa 和酰基辅酶 A：二酰基甘油酰基转移酶基因 aftAa，并表明通过失活它们，观察到细胞内葡萄糖-1-磷酸/葡萄糖-6-磷酸和丙二酰辅酶 A/乙酰辅酶 A 的浓度分别增加。每种单独的修饰都导致 MTM 产量增加，但通过组合所有策略，获得了最高的产量水平。此外，这些策略中的一些成功地应用于提高具有改善的药理学性质的四种 MTM 衍生物的产量：去甲酰基-密特霉素、去甲酰基-3D-β-D-地芰糖基-密特霉素、密特霉素 SK 和密特霉素 SDK。
