Deng Lingyi Lynn, Humphries Donald E, Arbeit Robert D, Carlton Laura E, Smole Sandra C, Carroll J David
Department of Medicine, Boston University School of Medicine, Research Service, VA Boston Healthcare System, 150 S. Huntington Ave., Boston, MA 02130, USA.
Biochim Biophys Acta. 2005 Feb 14;1747(1):57-66. doi: 10.1016/j.bbapap.2004.09.021. Epub 2004 Nov 11.
Mycobacterium tuberculosis is a major global pathogen whose threat has increased with the emergence of multidrug-resistant strains. The cell wall of M. tuberculosis is thick, rigid, and hydrophobic, which serves to protect the organism from the environment and makes it highly impermeable to conventional antimicrobial agents. There is little known about cell wall autolysins (also referred to as peptidoglycan hydrolases) of mycobacteria. We identified an open reading frame (Rv3915) in the M. tuberculosis genome designated cwlM that appeared consistent with a peptidoglycan hydrolase. The 1218-bp gene was amplified by PCR, cloned and expressed in E. coli strain HMS174(DE-3), and its gene product, a 47-kDa recombinant protein, was purified and partially characterized. Purified CwlM was able to lyse whole mycobacteria, release peptidoglycan from the cell wall of Micrococcus luteus and Mycobacterium smegmatis, and cleave N-acetylmuramoyl-L-alanyl-D-isoglutamine, releasing free N-acetylmuramic acid. These results indicate that CwlM is a novel autolysin and identify cwlM as the first, to our knowledge, autolysin gene identified and cloned from M. tuberculosis. CwlM offers a new target for a unique class of drugs that could alter the permeability of the mycobacterial cell wall and enhance the effectiveness of treatments for tuberculosis.
结核分枝杆菌是一种主要的全球病原体，随着多重耐药菌株的出现，其威胁日益增加。结核分枝杆菌的细胞壁厚、坚硬且疏水，这有助于保护该生物体免受外界环境影响，并使其对传统抗菌药物具有高度不渗透性。关于分枝杆菌的细胞壁自溶素（也称为肽聚糖水解酶），人们所知甚少。我们在结核分枝杆菌基因组中鉴定出一个开放阅读框（Rv3915），命名为cwlM，它似乎与一种肽聚糖水解酶一致。通过PCR扩增出1218个碱基对的基因，将其克隆并在大肠杆菌菌株HMS174(DE-3)中表达，其基因产物，一种47千道尔顿的重组蛋白，被纯化并进行了部分特性分析。纯化后的CwlM能够裂解完整的分枝杆菌，从藤黄微球菌和耻垢分枝杆菌的细胞壁释放肽聚糖，并切割N-乙酰胞壁酰-L-丙氨酰-D-异谷氨酰胺，释放出游离的N-乙酰胞壁酸。这些结果表明CwlM是一种新型自溶素，并据我们所知，将cwlM鉴定为首个从结核分枝杆菌中鉴定和克隆的自溶素基因。CwlM为一类独特的药物提供了一个新靶点，这类药物可以改变分枝杆菌细胞壁的通透性，提高结核病治疗的效果。
