Composition and Localization of Treponema denticola Outer Membrane Complexes
Composition and Localization of Treponema denticola Outer Membrane Complexes
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DOI：
10.1128/iai.05701-11
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发表时间：
2011-12-01
影响因子：
3.1
通讯作者：
Fenno, J. Christopher
中科院分区：
文献类型：
--
作者：
Godovikova, Valentina;Goetting-Minesky, M. Paula;Fenno, J. Christopher
齿垢密螺旋体外膜脂蛋白-蛋白酶复合物（牙本质素）通过降解细胞外基质成分和破坏细胞间宿主信号传导途径而导致牙周病。我们最近证明，prcB，位于上游和共转录与prcA和prtP，编码一个22 kDa的脂蛋白，与PrtP相互作用，并需要其活动。在这里，我们进一步表征产品的蛋白酶位点和它们的作用，表达，形成和定位的外膜复合物。PrcB在天然凝胶中作为包括PrtP和PrcA以及主要外鞘蛋白Msp的>400-kDa复合物的一部分迁移。PrcB可检测为纯化的活性蛋白酶复合物的次要成分，该复合物先前报道仅由PrtP和辅助多肽PrcA 1和PrcA 2组成。虽然它缺乏典型的核糖体结合位点存在上游的prcA和prtP，PrcB是目前在全细胞提取物中的PrtP的水平相似。免疫荧光显微镜显示PrtP、PrcA 1、PrcB和Msp成熟形式的细胞表面暴露。16-kDa的N-末端酰化片段的PrtP（预计将被释放在激活PrtP）存在于细胞提取物中，但既没有检测到在纯化的活性蛋白酶复合物，也没有在细胞表面上。PrcA 2，可检测到的Msp缺陷型细胞的表面上，但不是野生型细胞，与Msp共免疫沉淀。我们的研究结果表明，PrcB是外膜脂蛋白酶复合物的一个组成部分，Msp和PrcA 2的相互作用可能介导形成一个非常高分子量的外膜复合物。
The Treponema denticola outer membrane lipoprotein-protease complex (dentilisin) contributes to periodontal disease by degrading extracellular matrix components and disrupting intercellular host signaling pathways. We recently demonstrated that prcB, located upstream of and cotranscribed with prcA and prtP, encodes a 22-kDa lipoprotein that interacts with PrtP and is required for its activity. Here we further characterize products of the protease locus and their roles in expression, formation, and localization of outer membrane complexes. PrcB migrates in native gels as part of a >400-kDa complex that includes PrtP and PrcA, as well as the major outer sheath protein Msp. PrcB is detectable as a minor constituent of the purified active protease complex, which was previously reported to consist of only PrtP and auxiliary polypeptides PrcA1 and PrcA2. Though it lacks the canonical ribosome binding site present upstream of both prcA and prtP, PrcB is present at levels similar to those of PrtP in whole-cell extracts. Immunofluorescence microscopy demonstrated cell surface exposure of the mature forms of PrtP, PrcA1, PrcB, and Msp. The 16-kDa N-terminal acylated fragment of PrtP (predicted to be released during activation of PrtP) was present in cell extracts but was detected neither in the purified active protease complex nor on the cell surface. PrcA2, detectable on the surface of Msp-deficient cells but not that of wild-type cells, coimmunoprecipitated with Msp. Our results indicate that PrcB is a component of the outer membrane lipoprotein protease complex and that Msp and PrcA2 interaction may mediate formation of a very-high-molecular-weight outer membrane complex.