一种用于磷酸泛酰巯基乙胺基转移酶的均相共振能量转移测定法。
A homogeneous resonance energy transfer assay for phosphopantetheinyl transferase.
作者信息
Foley Timothy L, Burkart Michael D
机构信息
Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, CA 92093, USA.
出版信息
Anal Biochem. 2009 Nov 1;394(1):39-47. doi: 10.1016/j.ab.2009.06.037. Epub 2009 Jun 30.
Abstract
Phosphopantetheinyl transferase plays an essential role in activating fatty acid, polyketide, and nonribosomal peptide biosynthetic pathways, catalyzing covalent attachment of a 4'-phosphopantetheinyl group to a conserved residue within carrier protein domains. This enzyme has been validated as an essential gene to primary metabolism and presents a target for the identification of antibiotics with a new mode of action. Here we report the development of a homogeneous resonance energy transfer assay using fluorescent coenzyme A derivatives and a surrogate peptide substrate that can serve to identify inhibitors of this enzyme class. This assay lays a blueprint for translation of these techniques to other transferase enzymes that accept fluorescent substrate analogues.
摘要
磷酸泛酰巯基乙胺基转移酶在激活脂肪酸、聚酮化合物和非核糖体肽生物合成途径中发挥着至关重要的作用，它催化将一个4'-磷酸泛酰巯基乙胺基团共价连接到载体蛋白结构域内的一个保守残基上。该酶已被确认为初级代谢的必需基因，并成为鉴定具有新作用模式抗生素的一个靶点。在此，我们报告了一种使用荧光辅酶A衍生物和替代肽底物的均相共振能量转移测定法的开发，该方法可用于鉴定这类酶的抑制剂。该测定法为将这些技术应用于其他接受荧光底物类似物的转移酶奠定了基础。
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