A Comprehensive Study of the Interaction between Peptidoglycan Fragments and the Extracellular Domain of Mycobacterium tuberculosis Ser/Thr Kinase PknB.
A Comprehensive Study of the Interaction between Peptidoglycan Fragments and the Extracellular Domain of Mycobacterium tuberculosis Ser/Thr Kinase PknB.
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DOI：
10.1002/cbic.201700385
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发表时间：
2017-11-02
期刊：
影响因子：
--
通讯作者：
Fujimoto Y
中科院分区：
文献类型：
--
作者：
Wang Q;Marchetti R;Prisic S;Ishii K;Arai Y;Ohta I;Inuki S;Uchiyama S;Silipo A;Molinaro A;Husson RN;Fukase K;Fujimoto Y
结核分枝杆菌Ser/Thr激酶PknB参与细菌细胞生长和细胞分裂的调节。PknB的细胞内激酶功能被认为是由PknB的胞质外结构域识别的肽聚糖（PGN）片段触发的。在M.结核分枝杆菌具有几种不寻常的修饰，包括胞壁酸残基中存在N-羟乙酰基（除了N-乙酰基之外）和肽链中d-Glu的酰胺化。使用合成的PGN片段，将这些不同的PGN结构，我们分析了它们的结合特性，通过生物层干涉法（BLI），NMR光谱，和本地质谱（nMS）技术。BLI的结果表明，含有1，6-脱水MurNAc和较长聚糖链的肽具有较高的结合效力，并且肽茎的第四个氨基酸d-Ala对蛋白质识别至关重要。饱和转移差（STD）NMR光谱表明PASTA-PGN片段结合主要涉及茎肽区域。nMS表明结合化学计量比为1：1。这些数据为PGN与PknB的特异性相互作用提供了第一个分子基础，并牢固地确立了PGN作为PknB的有效配体。
The Mycobacterium tuberculosis Ser/Thr kinase PknB is implicated in the regulation of bacterial cell growth and cell division. The intracellular kinase function of PknB is thought to be triggered by peptidoglycan (PGN) fragments that are recognized by the extracytoplasmic domain of PknB. The PGN in the cell wall of M. tuberculosis has several unusual modifications, including the presence of N-glycolyl groups (in addition to N-acetyl groups) in the muramic acid residues and amidation of d-Glu in the peptide chains. Using synthetic PGN fragments incorporating these diverse PGN structures, we analyzed their binding characters through biolayer interferometry (BLI), NMR spectroscopy, and native mass spectrometry (nMS) techniques. The results of BLI showed that muropeptides containing 1,6-anhydro-MurNAc and longer glycan chains exhibited higher binding potency and that the fourth amino acid of the peptide stem, d-Ala, was crucial for protein recognition. Saturation transfer difference (STD) NMR spectroscopy indicated the major involvement of the stem peptide region in the PASTA-PGN fragment binding. nMS suggested that the binding stoichiometry was 1:1. The data provide the first molecular basis for the specific interaction of PGN with PknB and firmly establish PGNs as the effective ligands of PknB.
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