Department of Biochemistry and Microbiology, University of Victoria, Victoria, BC, Canada.
University of Victoria-Genome BC Proteomics Centre, University of Victoria, Victoria, BC, Canada.
Sci Rep. 2023 Oct 25;13(1):18259. doi: 10.1038/s41598-023-45219-8.
Comprehensive proteome-wide analysis of the syphilis spirochete, Treponema pallidum ssp. pallidum, is technically challenging due to high sample complexity, difficulties with obtaining sufficient quantities of bacteria for analysis, and the inherent fragility of the T. pallidum cell envelope which further complicates proteomic identification of rare T. pallidum outer membrane proteins (OMPs). The main aim of the present study was to gain a deeper understanding of the T. pallidum global proteome expression profile under infection conditions. This will corroborate and extend genome annotations, identify protein modifications that are unable to be predicted at the genomic or transcriptomic levels, and provide a foundational knowledge of the T. pallidum protein expression repertoire. Here we describe the optimization of a T. pallidum-specific sample preparation workflow and mass spectrometry-based proteomics pipeline which allowed for the detection of 77% of the T. pallidum protein repertoire under infection conditions. When combined with prior studies, this brings the overall coverage of the T. pallidum proteome to almost 90%. These investigations identified 27 known/predicted OMPs, including potential vaccine candidates, and detected expression of 11 potential OMPs under infection conditions for the first time. The optimized pipeline provides a robust and reproducible workflow for investigating T. pallidum protein expression during infection. Importantly, the combined results provide the deepest coverage of the T. pallidum proteome to date.
全面的梅毒螺旋体（苍白密螺旋体亚种）蛋白质组学分析在技术上具有挑战性，这是由于样品复杂性高、获得足够数量的细菌进行分析的困难以及梅毒螺旋体细胞包膜的固有脆弱性，这进一步使罕见的梅毒螺旋体外膜蛋白（OMP）的蛋白质组学鉴定复杂化。本研究的主要目的是更深入地了解感染条件下梅毒螺旋体的全蛋白质组表达谱。这将证实和扩展基因组注释，鉴定无法在基因组或转录组水平预测的蛋白质修饰，并为梅毒螺旋体蛋白质表达库提供基础知识。在这里，我们描述了梅毒螺旋体特异性样品制备工作流程和基于质谱的蛋白质组学管道的优化，该优化允许在感染条件下检测到 77%的梅毒螺旋体蛋白质组。当与先前的研究相结合时，这使梅毒螺旋体蛋白质组的整体覆盖率几乎达到 90%。这些研究鉴定了 27 种已知/预测的 OMP，包括潜在的疫苗候选物，并首次在感染条件下检测到 11 种潜在 OMP 的表达。优化的管道为研究感染期间梅毒螺旋体的蛋白质表达提供了一个稳健且可重复的工作流程。重要的是，综合结果提供了迄今为止对梅毒螺旋体蛋白质组的最深覆盖。
