Department of Biochemistry and Microbiology, University of Victoria, Victoria, British Columbia, Canada.
J Proteome Res. 2009 Oct;8(10):4564-78. doi: 10.1021/pr9004597.
Comparative global proteome analyses were performed on Leptospira interrogans serovar Copenhageni grown under conventional in vitro conditions and those mimicking in vivo conditions (iron limitation and serum presence). Proteomic analyses were conducted using iTRAQ and LC-ESI-tandem mass spectrometry complemented with two-dimensional gel electrophoresis and MALDI-TOF mass spectrometry. A total of 563 proteins were identified in this study. Altered expression of 65 proteins, including upregulation of the L. interrogans virulence factor Loa22 and 5 novel proteins with homology to virulence factors found in other pathogens, was observed between the comparative conditions. Immunoblot analyses confirmed upregulation of 5 of the known or putative virulence factors in L. interrogans exposed to the in vivo-like environmental conditions. Further, ELISA analyses using serum from patients with leptospirosis and immunofluorescence studies performed on liver sections derived from L. interrogans-infected hamsters verified expression of all but one of the identified proteins during infection. These studies, which represent the first documented comparative global proteome analysis of Leptospira, demonstrated proteome alterations under conditions that mimic in vivo infection and allowed for the identification of novel putative L. interrogans virulence factors.
进行了比较全蛋白质组分析，对在传统体外条件下和模拟体内条件（铁限制和血清存在）下生长的钩端螺旋体科 Copenhageni 进行了研究。使用 iTRAQ 和 LC-ESI-串联质谱联用二维凝胶电泳和 MALDI-TOF 质谱进行蛋白质组分析。在这项研究中总共鉴定了 563 种蛋白质。在比较条件下观察到 65 种蛋白质的表达发生变化，包括钩端螺旋体毒力因子 Loa22 的上调和 5 种与其他病原体毒力因子具有同源性的新型蛋白质。免疫印迹分析证实，在暴露于类似体内环境条件下的钩端螺旋体中，5 种已知或推测的毒力因子上调。此外，使用来自钩端螺旋体病患者的血清进行 ELISA 分析，并对来自感染钩端螺旋体的仓鼠的肝切片进行免疫荧光研究，证实了除一种外的所有鉴定蛋白在感染期间的表达。这些研究代表了首次对钩端螺旋体进行的比较全蛋白质组分析，证明了在模拟体内感染的条件下蛋白质组发生了变化，并确定了新的钩端螺旋体推测毒力因子。
