Virulence-associated subtilisin-like proteases that use a novel disulphide-tethered exosite to mediate substrate specificity (3LPA, 3LPC, 3LPD)

Many bacterial pathogens produce extracellular proteases that are involved in the degradation of the host extracellular matrix. Dichelobacter nodosus, which causes ovine footrot, is one such pathogen, Mutagenesis and virulence studies revealed that AprV2, one of three secreted subtilisin-like D. nodosus proteases, is required for virulence. Our work challenges the previous hypothesis that the elastase activity of AprV2 is important for disease progression, since aprV2 mutants were virulent when complemented with a variant with impaired elastase activity. These data reveal that an unusual extended disulphide-tethered loop functions as an exosite that governs the ability of AprV2 to degrade insoluble extracellular matrix components. The disulphide bond and Tyr92, located at the exposed end of the loop, were functionally important. Bioinformatics suggests that other pathogens utilize a similar mechanism, providing a new paradigm for understanding the role of proteases in disease.

众多细菌病原体分泌的胞外蛋白酶参与宿主胞外基质的降解。结节梭菌（Dichelobacter nodosus），引起羊蹄腐病的病原体之一，便属于此类。突变和致病性研究表明，AprV2，即三种分泌的类似Subtilisin的结节梭菌蛋白酶之一，对于致病性是必需的。我们的研究挑战了先前关于AprV2的弹性蛋白酶活性对于疾病进展重要的假设，因为当使用具有受损弹性蛋白酶活性的变体进行互补时，aprV2突变体仍表现出致病性。这些数据揭示了异常的延伸二硫键连接的环作为外位点，调控AprV2降解不溶性胞外基质成分的能力。二硫键和位于环暴露端的天冬酰胺92残基在功能上具有重要意义。生物信息学分析表明，其他病原体利用类似的机制，为理解蛋白酶在疾病中的作用提供了新的范式。