Kapur V, Topouzis S, Majesky M W, Li L L, Hamrick M R, Hamill R J, Patti J M, Musser J M
Department of Pathology, Baylor College of Medicine, Houston, Texas 77030.
Microb Pathog. 1993 Nov;15(5):327-46. doi: 10.1006/mpat.1993.1083.
Streptococcus pyogenes secretes an extracellular cysteine protease that cleaves human interleukin 1 beta precursor to form biologically active IL-1 beta, a major cytokine mediating inflammation and shock. To further investigate the potential role of the cysteine protease in host-parasite interactions, the enzyme was purified to apparent homogeneity and tested for ability to degrade several human extracellular matrix proteins. Purified protease cleaved fibronectin, apparently at specific sites, and rapidly degraded vitronectin. In contrast, the protease did not have substantial activity against laminin. The cysteine protease also cleaved fibronectin from human umbilical vein endothelial cells grown in vitro. Allelic variation in the cysteine protease structural gene was studied in 67 strains expressing 39 M protein serotypes and five provisional M serologic types, and representing 50 phylogenetically distinct clones identified by multilocus enzyme electrophoresis. The gene is well conserved and allelic variation is due solely to accumulation of point mutations. Based on predicted amino acid sequences, one mature cysteine protease variant would be made by clones expressing serotypes M2, M3, M4, M5, M6, M9, M10, M11, M12, M14, M18, M22, M23, M25, M27, M41, M49, M56, M59, two provisional M types, and two clones non-typeable for M protein. Moreover, 33 of the 39 speB alleles identified encode one of three mature protease variants that differ from one another at only one or two amino acids clustered in a ten-amino acid region. All 39 alleles, and virtually all strains, encode a product that reacts with polyclonal antisera specific for purified cysteine protease. No compelling evidence was found for a primitive differentiation of the speB gene into two distinct classes, as has been proposed for M protein, opacity factor phenotype, and vir regulon architecture. The results demonstrate that the cysteine protease is well conserved in natural populations of S. pyogenes, provide additional evidence that this enzyme is involved in host-parasite interactions, and suggest that the protease plays a role in bacterial dissemination, colonization, and invasion, and inhibition of wound healing.
化脓性链球菌分泌一种细胞外半胱氨酸蛋白酶，该酶可切割人白细胞介素1β前体以形成具有生物活性的IL-1β，IL-1β是介导炎症和休克的主要细胞因子。为了进一步研究半胱氨酸蛋白酶在宿主-寄生虫相互作用中的潜在作用，将该酶纯化至表观均一，并测试其降解几种人细胞外基质蛋白的能力。纯化的蛋白酶可切割纤连蛋白，显然是在特定位点，并能快速降解玻连蛋白。相比之下，该蛋白酶对层粘连蛋白没有显著活性。半胱氨酸蛋白酶还可切割体外培养的人脐静脉内皮细胞中的纤连蛋白。在表达39种M蛋白血清型和5种临时M血清型的67株菌株中研究了半胱氨酸蛋白酶结构基因的等位基因变异，这些菌株代表了通过多位点酶电泳鉴定的50个系统发育上不同的克隆。该基因高度保守，等位基因变异完全是由于点突变的积累。根据预测的氨基酸序列，表达血清型M2、M3、M4、M5、M6、M9、M10、M11、M12、M14、M18、M22、M23、M25、M27、M41、M49、M56、M59的克隆以及两种临时M型和两种无法分型的M蛋白克隆将产生一种成熟的半胱氨酸蛋白酶变体。此外，所鉴定的39个speB等位基因中的33个编码三种成熟蛋白酶变体之一，这三种变体彼此之间仅在聚集于一个十个氨基酸区域的一两个氨基酸上有所不同。所有39个等位基因以及几乎所有菌株都编码一种与针对纯化的半胱氨酸蛋白酶的多克隆抗血清发生反应的产物。没有发现令人信服的证据表明speB基因如对M蛋白、不透明因子表型和毒力调节子结构所提出的那样原始分化为两个不同的类别。结果表明，半胱氨酸蛋白酶在化脓性链球菌的自然群体中高度保守，提供了该酶参与宿主-寄生虫相互作用的额外证据，并表明该蛋白酶在细菌传播、定植和侵袭以及抑制伤口愈合中起作用。
