In vitro peptidoglycan synthesis assay with lipid II substrate.
In vitro peptidoglycan synthesis assay with lipid II substrate.
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DOI：
10.1007/978-1-62703-245-2_17
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发表时间：
2013-01-01
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影响因子：
--
通讯作者：
Vollmer, Waldemar
中科院分区：
文献类型：
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作者：
Biboy, Jacob;Bui, Nhat Khai;Vollmer, Waldemar
细菌细胞壁肽聚糖是由脂质II前体通过两个反应合成的。糖基转移酶聚合聚糖链，转肽酶形成肽交联。双功能 A 类青霉素结合蛋白催化这两种反应。在这里，我们描述了一种体外肽聚糖合成测定，利用放射性标记的脂质 II 底物来同时监测肽聚糖糖基转移酶和转肽酶活性。通过高压液相色谱分离反应产物并通过流通闪烁计数进行定量。
Bacterial cell wall peptidoglycan is synthesized from lipid II precursor by two reactions. Glycosyltransferases polymerize the glycan chains and transpeptidases form the peptide cross-links. The bifunctional class A penicillin-binding proteins catalyze both of these reactions. Here, we describe an in vitro peptidoglycan synthesis assay utilizing radiolabeled lipid II substrate to monitor simultaneously peptidoglycan glycosyltransferase and transpeptidase activities. The products of the reaction are separated by high-pressure liquid chromatography and quantified by flow-through scintillation counting.