Shaw Julie L V, Smith Christopher R, Diamandis Eleftherios P
Department of Pathology and Laboratory Medicine, Mount Sinai Hospital, Toronto, Ontario, Canada.
J Proteome Res. 2007 Jul;6(7):2859-65. doi: 10.1021/pr0701658. Epub 2007 Jun 14.
Human cervico-vaginal fluid (CVF) is a mixture of fluids originating from the vagina, cervix, endometrium, and oviduct. CVF has been shown to play an important role in protecting the vagina from infection. We used "bottom-up" proteomic approaches to characterize the protein repertoire of human CVF. We applied two different sample prefractionation methods, one-dimensional-SDS-PAGE (1D-SDS-PAGE) and strong cation-exchange chromatography, followed by LC-MS/MS and bioinformatic analysis. We identified a total of 685 proteins. Strong cation-exchange chromatography prefractionation resulted in a larger number of proteins identified when compared with 1D-SDS-PAGE. Extracellular or membrane proteins made up 30% of the proteins identified, according to Genome Ontology (GO) classifications. We confirmed the presence of defense-related proteins, such as haptoglobin, defensins, and lactoferrin; and identified new ones such as azurocidin and dermcidin. We also identified many serine and cysteine proteases, including 6 members of the kallikrein family (KLKs 6, 7, 10, 11, 12, and 13). The same KLKs were also confirmed quantitatively by ELISA assays. Knowledge of the CVF proteome will aid in the discovery of potential biomarkers for gynecological malignancies and infections and provide additional clues for its physiological functions.
人宫颈阴道液(CVF)是源自阴道、子宫颈、子宫内膜和输卵管的多种液体的混合物。CVF已被证明在保护阴道免受感染方面发挥重要作用。我们采用“自下而上”的蛋白质组学方法来表征人CVF的蛋白质组成。我们应用了两种不同的样品预分级方法,一维SDS-PAGE(1D-SDS-PAGE)和强阳离子交换色谱法,随后进行液相色谱-串联质谱(LC-MS/MS)和生物信息学分析。我们总共鉴定出685种蛋白质。与1D-SDS-PAGE相比,强阳离子交换色谱预分级鉴定出的蛋白质数量更多。根据基因本体论(GO)分类,细胞外或膜蛋白占鉴定出的蛋白质的30%。我们证实了防御相关蛋白的存在,如触珠蛋白、防御素和乳铁蛋白;并鉴定出了新的蛋白,如天青杀素和皮肤抗菌肽。我们还鉴定出许多丝氨酸和半胱氨酸蛋白酶,包括激肽释放酶家族(KLKs 6、7、10、11、12和13)的6个成员。通过酶联免疫吸附测定(ELISA)对相同的KLKs进行了定量确认。了解CVF蛋白质组将有助于发现妇科恶性肿瘤和感染的潜在生物标志物,并为其生理功能提供更多线索。