Post-translational modifications (PTMs) of surface-glycoprotein GP82, the major adhesin on metacyclic forms of Trypanosoma cruzi

• Grant number: 12/14369-3
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• Start date: February 01, 2013
• End date: January 31, 2015
• Field of knowledge: Biological Sciences - Parasitology - Protozoology of Parasites
• Principal Investigator: José Franco da Silveira Filho
• Grantee: Esteban Mauricio Cordero Veas
• Host Institution: Escola Paulista de Medicina (EPM). Universidade Federal de São Paulo (UNIFESP). Campus São Paulo. São Paulo , SP, Brazil

Abstract

Post-translational modifications (PTMs) are responsible for modifications in size, composition, function and/or localization of proteins. Among those, the glycosylphosphatidylinositol anchor-addition alters the size and composition of proteins, by removing the amino- and carboxy-terminus. This PTM, conserved among eukaryotes, is a co-translational process due in the endoplasmic reticulum. Here the signal peptidase removes the amino-terminus from the nascent protein and the GPI-transamidase catalyzes the cleavage/addition of the GPI-anchor moiety in the carboxy-terminus.The surface glycoprotein GP82 from infective metacyclic forms of Trypanosoma cruzi is a stage-specific adhesin attached to the parasite surface's by a GPI anchor. The GP82 is actively involved in the parasite's adhesion/invasion to the mammalian host cells. In silico analyses have predicted the putative cleavage sites at the amino- and carboxy-terminus, but there is no experimental confirmation yet.In order to characterize the amino- and carboxy-terminus derived from the mature GP82 protein, we intend to transfect the parasite T. cruzi with several constructs expressing the GP82, either tagged with the c-myc epitope or in frame with the GFP. The tagged proteins will be purified by affinity chromatography and submitted to chemical and/or enzymatic proteolysis. The resulting peptides will be analyzed by Edman's degradation and mass spectrometry. The mass spectrometry is due to overcome the difficulties found in the Edman technique when the peptides have their amino-terminus blocked.Several members of the TS superfamily and others GPI-anchored proteins families of T. cruzi such as GP63, MAP and mucins, share a significant degree of conservation at the amino- and carboxy-terminus. As the signals involved in the GPI-anchoring are mainly located at the amino- and carboxy-terminus, the protocols herein described to analyze the GP82 glycoprotein could be applied to the study of another TS superfamily members along with others T. cruzi GPI-anchored protein families.

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

CORDERO, ESTEBAN M.; CORTEZ, CRISTIAN; YOSHIDA, NOBUKO; DA SILVEIRA, JOSE FRANCO. Signal peptide recognition in Trypanosoma cruzi GP82 adhesin relies on its localization at protein N-terminus. SCIENTIFIC REPORTS, v. 9, MAY 13 2019. (12/14369-3, 11/51475-3)