Departments of Medicine, University of Connecticut Health Center, Farmington, Connecticut, USA.
J Bacteriol. 2012 May;194(9):2321-33. doi: 10.1128/JB.00101-12. Epub 2012 Mar 2.
Identification of Treponema pallidum rare outer membrane proteins (OMPs) has been a longstanding objective of syphilis researchers. We recently developed a consensus computational framework that employs a battery of cellular localization and topological prediction tools to generate ranked clusters of candidate rare OMPs (D. L. Cox et al., Infect. Immun. 78:5178-5194, 2010). TP0117/TP0131 (TprC/D), a member of the T. pallidum repeat (Tpr) family, was a highly ranked candidate. Circular dichroism, heat modifiability by SDS-PAGE, Triton X-114 phase partitioning, and liposome incorporation confirmed that full-length, recombinant TprC (TprC(Fl)) forms a β-barrel capable of integrating into lipid bilayers. Moreover, TprC(Fl) increased efflux of terbium-dipicolinic acid complex from large unilamellar vesicles and migrated as a trimer by blue-native PAGE. We found that in T. pallidum, TprC is heat modifiable, trimeric, expressed in low abundance, and, based on proteinase K accessibility and opsonophagocytosis assays, surface exposed. From these collective data, we conclude that TprC is a bona fide rare OMP as well as a functional ortholog of Escherichia coli OmpF. We also discovered that TprC has a bipartite architecture consisting of a soluble N-terminal portion (TprC(N)), presumably periplasmic and bound directly or indirectly to peptidoglycan, and a C-terminal β-barrel (TprC(C)). Syphilitic rabbits generate antibodies exclusively against TprC(C), while secondary syphilis patients fail to mount a detectable antibody response against either domain. The syphilis spirochete appears to have resolved a fundamental dilemma arising from its extracellular lifestyle, namely, how to enhance OM permeability without increasing its vulnerability to the antibody-mediated defenses of its natural human host.
鉴定苍白密螺旋体的罕见外膜蛋白(OMPs)一直是梅毒研究人员的长期目标。我们最近开发了一种共识计算框架,该框架使用一系列细胞定位和拓扑预测工具,生成候选罕见 OMP 的排序聚类(D. L. Cox 等人,Infect. Immun. 78:5178-5194, 2010)。TP0117/TP0131(TprC/D)是苍白密螺旋体重复(Tpr)家族的成员,是一个排名很高的候选者。圆二色性、SDS-PAGE 的热可变性、Triton X-114 相分离和脂质体掺入证实全长重组 TprC(TprC(Fl))形成能够整合到脂质双层中的β-桶。此外,TprC(Fl)增加了从大单室囊泡中铽-二吡啶酸络合物的外排,并且通过蓝-native PAGE 迁移为三聚体。我们发现,在苍白密螺旋体中,TprC 可热修饰、三聚体、低丰度表达,并且根据蛋白水解酶可及性和调理吞噬作用测定,表面暴露。从这些综合数据中,我们得出结论,TprC 是一种真正的罕见 OMP,也是大肠杆菌 OmpF 的功能同源物。我们还发现 TprC 具有由可溶性 N 端部分(TprC(N))组成的二部分结构,推测是周质,直接或间接与肽聚糖结合,以及 C 端β-桶(TprC(C))。梅毒兔仅产生针对 TprC(C)的抗体,而二期梅毒患者未能对任一结构域产生可检测的抗体反应。梅毒螺旋体似乎解决了其体外生活方式所带来的一个根本困境,即如何在不增加其对天然人类宿主抗体防御的脆弱性的情况下提高 OM 通透性。