α,β → β,γ double bond migration in corallopyronin A biosynthesis
α,β → β,γ double bond migration in corallopyronin A biosynthesis
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DOI：
10.1039/c3sc51854j
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发表时间：
2013-01-01
期刊：
影响因子：
8.4
通讯作者：
Schaeberle, Till F.
中科院分区：
文献类型：
--
作者：
Lohr, Friederike;Jenniches, Imke;Schaeberle, Till F.
在聚酮生物合成中，将-羰基还原成烯烃通常会产生-、-双键。然而，在一些抗生素中，在β、γ位置观察到罕见的碳-碳双键。体内活性抗生素corallopyronin A代表了这样一种分子，在分子组装过程中发生了α， β -> β， γ双键迁移。在这里，我们报告了负责这种双键异构化的酶结构域的体外分析。这种“移位域”是异源表达的，并与酰基载体蛋白结合的底物进行了测定。为了便于分析，生物合成的corallopyronin A中间体被化学合成为snac衍生物。酶活性通过核磁共振和高分辨率质谱测量进行分析，后者通过在氘化缓冲液中进行分析而启用。突变的酶变体提供了第一个实验证据必需氨基酸参与双键迁移。这些结果进一步支持了珊瑚pyronin A的生物合成。
In polyketide biosynthesis the reduction of beta-carbonyl groups to an alkene usually results in a alpha,beta double bond. However, in a few antibiotics the rare case of such a carbon-carbon double bond in beta,gamma position is observed. The in vivo active antibiotic corallopyronin A represents such a molecule, whereby a alpha,beta -> beta,gamma double bond migration takes place during the assembly of the molecule. Here we report the in vitro analysis of the enzyme domain responsible for this double bond isomerization. This "shift domain" was heterologously expressed and assayed with its acyl carrier protein bound substrate. To facilitate this analysis the biosynthetic corallopyronin A intermediate was chemically synthesized as a SNAC-derivative. Enzyme activity was analyzed by NMR and high-resolution MS measurements, the latter enabled by performing the assay in deuterated buffer. Mutated enzyme variants gave first experimental evidence for the essential amino acids involved in double bond migration. These results further support the proposed corallopyronin A biosynthesis.