Division of Rheumatology, Immunology, and Allergy, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.
Mass Spectrometry Resource, Division of Endocrinology, Metabolism, and Lipid Research, Department of Medicine, Washington University School of Medicine, St. Louis, MO, USA.
Methods Mol Biol. 2021;2306:227-238. doi: 10.1007/978-1-0716-1410-5_15.
This book chapter provides readers the step-by-step instruction for cell growth, lipid isolation, and lipid analysis to obtain the lipidome of Corynebacterium glutamicum (C. glutamicum) in the genus Corynebacterium, a biotechnologically important bacterium. We separate the lipid families by preparative HPLC with an analytical C-8 column, followed by linear ion-trap multiple stage mass spectrometry (LIT MS) with high-resolution mass measurement to define the structures of cytidine diphosphate diacylglycerol (CDP-DAG), glucuronosyl diacylglycerol (GlcA-DAG), α-D-mannopyranosyl-(1 → 4)-α-D-glucuronyl diacylglycerol (Man-GlcA-DAG), 1-mycolyl-2-acyl-phosphatidylglycerol (MA-PG), and acyl trehalose monomycolate (acyl-TMM) whose structures have been previously mis-assigned or not defined by mass spectrometric means. We also define the structures of mycolic acid, phosphatidylglycerol, phosphatidylinositol, cardiolipin, trehalose dimycolate lipids in the cell wall. The similarity of the lipidome to that in the Mycobacterium genera is consistent with the notion that Corynebacterium and Mycobacterium are gram-positive bacteria belonging to the suborder Corynebacterineae.
本章为读者提供了从细胞生长、脂质分离到脂质组分析的分步说明，以获得棒状杆菌属（Corynebacterium）中谷氨酸棒状杆菌（Corynebacterium glutamicum）的脂质组。我们使用制备型 HPLC 和分析型 C-8 柱分离脂质家族，然后进行线性离子阱多级质谱（LIT MS）分析，采用高分辨率质量测量方法来确定胞苷二磷酸二酰基甘油（CDP-DAG）、葡萄糖醛酸二酰基甘油（GlcA-DAG）、α-D-甘露吡喃基-(1→4)-α-D-葡萄糖醛酸二酰基甘油（Man-GlcA-DAG）、1-霉酚酰基-2-酰基-磷脂酰甘油（MA-PG）和酰基海藻糖单酯（acyl-TMM）的结构，这些结构之前通过质谱手段被错误分配或未被定义。我们还确定了细胞壁中分枝菌酸、磷脂酰甘油、磷脂酰肌醇、心磷脂和海藻糖二酯脂质的结构。脂质组与分枝杆菌属的脂质组相似，这与棒状杆菌属和分枝杆菌属均为革兰氏阳性菌、属于棒状杆菌亚目这一概念一致。
