Haydock Stephen F, Appleyard Anthony N, Mironenko Tatiana, Lester John, Scott Natasha, Leadlay Peter F
Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, UK.
Microbiology (Reading). 2005 Oct;151(Pt 10):3161-3169. doi: 10.1099/mic.0.28194-0.
The macrolide antibiotic concanamycin A has been identified as an exceptionally potent inhibitor of the vacuolar (V-type) ATPase. Such compounds have been mooted as the basis of a potential drug treatment for osteoporosis, since the V-ATPase is involved in the osteoclast-mediated bone resorption that underlies this common condition. To enable combinatorial engineering of altered concanamycins, the biosynthetic gene cluster governing the biosynthesis of concanamycin A has been cloned from Streptomyces neyagawaensis and shown to span a region of over 100 kbp of contiguous DNA. An efficient transformation system has been developed for S. neyagawaensis and used to demonstrate the role of the cloned locus in the formation of concanamycin A. Sequence analysis of the 28 ORFs in the region has revealed key features of the biosynthetic pathway, in particular the biosynthetic origin of portions of the backbone, which arise from the unusual polyketide building blocks ethylmalonyl-CoA and methoxymalonyl-ACP, and the origin of the pendant deoxysugar moiety 4'-O-carbamoyl-2'-deoxyrhamnose, as well as the presence of a modular polyketide synthase (PKS) encoded by six giant ORFs. Examination of the methoxymalonyl-specific acyltransferase (AT) domains has led to recognition of an amino acid sequence motif which can be used to distinguish methylmalonyl-CoA- from methoxymalonyl-ACP-specific AT domains in natural PKSs.
大环内酯类抗生素 concanamycin A 已被确认为液泡型（V 型）ATP 酶的一种极其有效的抑制剂。由于 V-ATP 酶参与破骨细胞介导的骨吸收过程，而这种骨吸收是骨质疏松这种常见病症的基础，因此这类化合物已被视为潜在骨质疏松症药物治疗的基础。为了实现对改造后的 concanamycins 进行组合工程改造，已从根岸链霉菌中克隆出控制 concanamycin A 生物合成的生物合成基因簇，该基因簇跨度超过 100 kbp 的连续 DNA 区域。已为根岸链霉菌开发了一种高效转化系统，并用于证明克隆位点在 concanamycin A 形成中的作用。对该区域 28 个开放阅读框的序列分析揭示了生物合成途径的关键特征，特别是主链部分的生物合成起源，其源于不寻常的聚酮化合物构建单元乙基丙二酰辅酶 A 和甲氧基丙二酰 - ACP，以及侧链脱氧糖部分 4'-O-氨基甲酰基-2'-脱氧鼠李糖的起源，同时还发现了由六个巨大开放阅读框编码的模块化聚酮合酶（PKS）。对甲氧基丙二酰特异性酰基转移酶（AT）结构域的研究导致识别出一种氨基酸序列基序，该基序可用于区分天然 PKS 中甲基丙二酰辅酶 A 特异性和甲氧基丙二酰 - ACP 特异性的 AT 结构域。
