Identification and characterization of the immunogenic cytotoxic TvCP39 proteinase gene of Trichomonas vaginalis

. Using a proteomic approach (A–C), the TvCP39 proteinase was identified as a papain-like proteinases encoded by a 915-bp cathepsin L-like cysteine proteinase gene ( tvcp39), which was amplified using PCR, cloned, and sequenced. A recombinant protein fragment corresponding to the mature region (TvCP39r) bound to the surface of HeLa cells (D–F) and protected them from trichomonal destruction (G). In addition, TvCP39 was found in vaginal secretions of patients with trichomonosis using an anti-TvCP39r antibody. [Display omitted] ► The TvCP39 proteinase of Trichomonas vaginalis was identified using a proteomic approach. ► TvCP39 is encoded by a 915-bp cathepsin L-like cysteine proteinase gene ( tvcp39). ► A recombinant TvCP39 mature fragment (TvCP39r) binds to the surface of HeLa cells and protects them from trichomonal destruction. ► TvCP39 is a glycosylated cysteine proteinase. ► The anti-TvCP39r antibody detects TvCP39 in vaginal secretions of patients with trichomonosis. ► TvCP39 is a potential biomarker for trichomonosis. TvCP39 is a 39 kDa cysteine proteinase (CP) involved in Trichomonas vaginalis cytotoxicity that has been found in vaginal secretions and is immunogenic in patients with trichomonosis. The goal of this work was to identify, clone, express, and characterize the tvcp39 gene. The tvcp39 gene was identified using a proteomic approach, and the complete gene was amplified using PCR, cloned, and sequenced. TvCP39 is encoded by a 915-bp cathepsin L-like CP gene. A fragment corresponding to the mature region (TvCP39r) was expressed, purified, and used to produce rabbit polyclonal antibodies and in functional assays. In one- and two-dimensional western blot assays, the anti-TvCP39r antibody reacted with two protein bands of ∼28 and 27 kDa and three spots of ∼28, 27, and 24 kDa in trichomonad proteinase-rich extracts that could correspond to the mature and processed fragments of the TvCP39 peptidase. The anti-TvCP39r antibody reacted with the parasitic surface and the native TvCP39 present in vaginal washes from patients with trichomonosis. Moreover, the recombinant TvCP39 protein bound to the surface of HeLa cells and protected HeLa cell monolayers from trichomonal destruction in a concentration-dependent manner. In conclusion, our data support TvCP39 as one of the surface proteinases that is glycosylated and is involved in trichomonal cytotoxicity. Thus, TvCP39 is the first glycosylated cysteine proteinase detected in T. vaginalis.