Institute of Biostructures and Bioimaging, CNR, Via Mezzocannone 16, I-80134, Naples, Italy.
Structure. 2010 Sep 8;18(9):1184-90. doi: 10.1016/j.str.2010.06.007.
Cell separation depends on cell-wall hydrolases that cleave the peptidoglycan layer connecting daughter cells. In Mycobacterium tuberculosis, this process is governed by the predicted endopeptidase RipA. In the absence of this enzyme, the bacterium is unable to divide and exhibits an abnormal phenotype. We here report the crystal structure of a relevant portion of RipA, containing its catalytic-domain and an extra-domain of hitherto unknown function. The structure clearly demonstrates that RipA is produced as a zymogen, which needs to be activated to achieve cell-division. Bacterial cell-wall degradation assays and proteolysis experiments strongly suggest that activation occurs via proteolytic processing of a fully solvent exposed loop identified in the crystal structure. Indeed, proteolytic cleavage at this loop produces an activated form, consisting of the sole catalytic domain. Our work provides the first evidence of self-inhibition in cell-disconnecting enzymes and opens a field for the design of novel antitubercular therapeutics.
细胞分离依赖于能够切割连接子细胞的肽聚糖层的细胞壁水解酶。在结核分枝杆菌中，这个过程由预测的内切肽酶 RipA 控制。在没有这种酶的情况下，细菌无法分裂，表现出异常的表型。我们在此报告了 RipA 的相关部分的晶体结构，其中包含其催化结构域和一个以前未知功能的额外结构域。该结构清楚地表明，RipA 作为酶原产生，需要激活才能实现细胞分裂。细菌细胞壁降解测定和蛋白水解实验强烈表明，激活是通过在晶体结构中鉴定的完全暴露于溶剂的环的蛋白水解处理来实现的。事实上，在该环处的蛋白水解切割产生了一种激活形式，由唯一的催化结构域组成。我们的工作提供了自抑制在细胞分离酶中的第一个证据，并为设计新型抗结核治疗药物开辟了一个领域。
