Trypanosoma cruzi serinecarboxipeptidase is a sulfated glycoprotein and a minor antigen in human Chagas disease infection
In this work, the presence of sulfated N-glycans was studied in a high-mannose-type glycoprotein of Trypanosoma cruzi with serinecarboxipeptidase (TcSCP) activity. The immune cross-reactivity between purified SCP and Cruzipain (Cz) was evidenced using rabbit sera specific for both glycoproteins. Taking advantage that SCP co-purifies with Cz from Concanavalin-A affinity columns, the Cz–SCP mixture was desulfated, ascribing the cross-reactivity to the presence of sulfate groups in both molecules. Therefore, knowing that Cz is a sulfated glycoprotein, with antigenic sulfated epitopes (sulfotopes), SCP was excised from SDS-PAGE and the N-glycosydic chains were analyzed by UV–MALDI–TOF-MS, confirming the presence of short-sulfated high-mannose-type oligosaccharidic chains. Besides, the presence of sulfotopes was analyzed in lysates of the different parasite stages demonstrating that a band with apparent molecular weight similar to SCP was highly recognized in trypomastigotes. In addition, SCP was confronted with sera of infected people with different degrees of cardiac dysfunction. Although most sera recognized it in different groups, no statistical association was found between sera antibodies specific for SCP and the severity of the disease. In summary, our findings demonstrate (1) the presence of sulfate groups in the N-glycosidic short chains of native TcSCP, (2) the existence of immune cross-reactivity between Cz and SCP, purified from epimastigotes, (3) the presence of common sulfotopes between both parasite glycoproteins, and (4) the enhanced presence of sulfotopes in trypomastigotes, probably involved in parasite–host relationship and/or infection. Interestingly, we show for the first time that SCP is a minor antigen recognized by most of chronic Chagas disease patient’s sera.
KeywordsTrypanosoma cruzi Serinecarboxypeptidase Glycomics Sulfate groups Sulfotopes Chagas’ disease Antigenicity
The authors thank to the Consejo Nacional de Investigaciones Científicas y Técnicas, CONICET, (PIP 11220110100660; PIP 07912012-2014), Universidad de Buenos Aires, UBA, (20020130100476BA) and Agencia Nacional de Promoción Científica y Tecnológica (ANPCyT) PICT-2013-0736 grants as well as to the Instituto Nacional de Parasitología “Dr M. Fatala Chaben”, ANLIS-Malbrán, Ministerio de Salud de la Nación, Argentina. The Ultraflex II (Bruker) TOF/TOF mass spectrometer was supported by the ANPCyT, (Grant PME 125, CEQUIBIEM).
- 1.World Health Organization (2014) WHO Fact Sheet No 340. http://www.who.int/mediacentre/factsheets/fs340/en/. Accessed 29 May 2014
- 12.Bastos IM, Grellier P, Martins NF, Cadavid-Restrepo G, de Souza-Ault MR, Augustyns K, Teixeira AR, Schrevel J, Maigret B, da Silveira JF, Santana JM (2005) Molecular, functional and structural properties of the prolyl oligopeptidase of Trypanosoma cruzi (POP Tc80), which is required for parasite entry into mammalian cells. Biochem J 388:29–38CrossRefPubMedCentralPubMedGoogle Scholar
- 17.Barrett A, Rawlings N, Woessner J (2004) Handbook of proteolytic enzymes. Elsevier Academic Press, LondonGoogle Scholar
- 19.Ferreira KA, Fajardo EF, Baptista RP, Macedo AM, Lages-Silva E, Ramírez LE, Pedrosa AL (2014) Species-specific markers for the differential diagnosis of Trypanosoma cruzi and Trypanosoma rangeli and polymorphisms detection in Trypanosoma rangeli. Parasitol Res 113(6):2199–2207CrossRefPubMedGoogle Scholar
- 21.Kawasaki N, Ohta M, Hyuga S, Hyuga M, Hayakawa T (2000) Application of liquid chromatography mass spectrometry and liquid chromatography with tandem mass spectrometry to the analysis of the site specific carbohydrate heterogeneity in erythropoietin. Anal Biochem 285:82–91CrossRefPubMedGoogle Scholar
- 37.World Health Organization (1991) Control of Chagas disease. Report of a WHO Expert Committee. World Health Organ Tech Rep Ser 811:1–95Google Scholar