Genetic manipulation of the biosynthetic process leading to phoslactomycins, potent protein phosphatase 2A inhibitors
Genetic manipulation of the biosynthetic process leading to phoslactomycins, potent protein phosphatase 2A inhibitors
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DOI：
10.1007/s10295-006-0116-1
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发表时间：
2006-07-01
影响因子：
3.4
通讯作者：
Reynolds, Kevin
中科院分区：
文献类型：
--
作者：
Ghatge, Mohini;Palaniappan, Nadaraj;Reynolds, Kevin
磷乳菌素(PLM)是由多种链霉菌分泌的一类不寻常的天然产物，含有α，β-不饱和三角洲内酯、氨基、磷酸酯、共轭二烯和环己烷环。磷霉素类、磷脂类和亮霉素类都含有相同的结构基团，只是C-18位的酰基取代基有所不同。这些化合物具有抗真菌或抗肿瘤活性，或两者兼有。PLM类化合物的抗肿瘤活性归因于对蛋白磷酸酶2A(PP2A)的有效和选择性抑制。PP2Ac-亚基的半胱氨酸-269位残基是PLM共价修饰的部位。在本文中，我们回顾了以前关于隔离的工作。PLM及其相关化合物的结构阐明和生物活性，以及我们在PLM稳定性和生物合成过程的遗传操作方面的工作现状。我们的工作表明PLM B在溶液中具有惊人的稳定性，最适pH为6。利用同位素标记的莽草酸和环己烷羧酸(CHC)进行的初步生物合成研究表明，PLM B是一种以CHC为起始单元合成的聚酮类抗生素。以柯林链霉菌CHC-CoA生物合成基因中的ChcA基因为探针，在链霉菌基因组中定位了一个75kb的29个开放阅读框编码PLM生物合成的区域。菌株HK803。对基因簇中的phns(2)和phnR(2)的分析和随后的操作允许对只产生一种PLM类似物的菌株进行合理的工程。PLM B，滴度比野生型高9倍。还产生了一株产生PLM G(PLM生物合成的倒数第二个中间体)的菌株。目前的工作是在体外选择性地将PLM G与各种羧酸进行酰化反应，并在CHCA缺失突变体中进行前体导向的生物合成，目的是产生新的PLM类似物。
Phoslactomycins (PLMs) represent an unusual structural class of natural products secreted by various streptomycetes, containing an alpha,beta-unsaturated delta-lactone, an amino group, phosphate ester, conjugated diene and a cyclohexane ring. Phosphazomycins, phospholines and leustroducsins contain the same structural moieties, varying only in the acyl substituent at the C-18 hydroxyl position. These compounds possess either antifungal or antitumor activities or both. The antitumor activity of the PLM class of compounds has been attributed to a potent and selective inhibition of protein phosphatase 2A (PP2A). The cysteine-269 residue of PP2Ac-subunit has been shown to be the site of covalent modification by PLMs. In this article, we review previous work on the isolation. structure elucidation and biological activities of PLMs and related compounds and current status of our work on both PLM stability and genetic manipulation of the biosynthetic process. Our work has shown that PLM B is surprisingly stable in solution, with a pH optimum of 6. Preliminary biosynthetic studies utilizing isotopically labeled shikimic acid and cyclohexanecarboxylic acid (CHC) suggested PLM B to be a polyketide-type antibiotic synthesized using CHC as a starter unit. Using a gene (chcA) from a set of CHC-CoA biosynthesis genes from Streptomyces collinits as a probe, a 75 kb region of 29 ORFs encoding PLM biosynthesis was located in the genome of Streptomyces sp. strain HK803. Analysis and subsequent manipulation of phnS(2) and phnR(2) in the gene cluster has allowed for rational engineering of a strain that produces only one PLM analog. PLM B, at ninefold higher titers than the wild type strain. A strain producing PLM G (the penultimate intermediate in PLMs biosynthesis) has also been generated. Current work is aimed at selective in vitro acylation of PLM G with various carboxylic acids and a precursor-directed biosynthesis in a chcA deletion mutant with the aim of generating novel PLM analogs.