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Thermo Savant FastPrep 120 Cell Disrupter System

公司名称: GMI
产品编号: 8357-30-1323
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Analysis of N-acetylmuramic acid-6-phosphate (MurNAc-6P) Accumulation by HPLC-MS
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2017-08-05
[Abstract]  We describe here in detail a high-performance liquid chromatography-mass spectrometry (HPLC-MS)-based method to determine N-acetylmuramic acid-6-phosphate (MurNAc-6P) in bacterial cell extracts. The method can be applied to both Gram-negative and Gram-positive bacteria, and as an example we use Escherichia coli cells in this study. Wild type and mutant cells are grown for a defined time in a medium of choice and harvested by centrifugation. Then the cells are disintegrated and soluble cell extracts are generated. After removal of proteins by precipitation with acetone, the ... [摘要]  我们在这里详细描述了一种基于高效液相色谱 - 质谱(HPLC-MS)的方法,以确定细菌细胞提取物中的N-乙酰谷氨酸-6-磷酸(MurNAc-6P)。该方法可以应用于革兰氏阴性菌和革兰氏阳性菌,作为一个例子,我们在本研究中使用大肠杆菌细胞。野生型和突变型细胞在选择的培养基中生长一定时间并通过离心收获。然后细胞分解并产生可溶性细胞提取物。通过用丙酮沉淀除去蛋白质后,通过HPLC-MS分析提取物。野生型和突变型细胞提取物的基本峰图谱用于测定显示两种样品中MurNAc-6P含量差异的差异离子谱。确定阴离子模式下MurNAc-6P((MH)= 372.070m/z的提取色谱图的峰面积允许定量使用的MurNAc-6P水平计算消泡聚糖的MurNAc含量的回收率。
【背景】在细菌生长期间,大部分细菌的肽聚糖细胞壁稳定地翻转并可能回收(再循环)。 肽聚糖回收代谢的关键化合物是积聚在大肠杆菌的MurNAc-6P醚酶(MurQ)突变体中的N-乙酰谷氨酸-6-磷酸(MurNAc-6P) (Jaeger等人,2005; Uehara等人,2006)。 MurQ直向同源物被发现在许多细菌中,包括革兰氏阳性菌(Litzinger等人,2010; Reith和Mayer,2011)。 ...

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