title: The Trypanosoma cruzi virulence factor oligopeptidase B (OPBTc) assembles into an active and stable dimer.
abstract: Oligopeptidase B, a processing enzyme of the prolyl oligopeptidase family, is considered as an important virulence factor in trypanosomiasis. Trypanosoma cruzi oligopeptidase B (OPBTc) is involved in host cell invasion by generating a Ca(2+)-agonist necessary for recruitment and fusion of host lysosomes at the site of parasite attachment. The underlying mechanism remains unknown and further structural and functional characterization of OPBTc may help clarify its physiological function and lead to the development of new therapeutic molecules to treat Chagas disease. In the present work, size exclusion chromatography and analytical ultracentrifugation experiments demonstrate that OPBTc is a dimer in solution, an association salt and pH-resistant and independent of intermolecular disulfide bonds. The enzyme retains its dimeric structure and is fully active up to 42°C. OPBTc is inactivated and its tertiary, but not secondary, structure is disrupted at higher temperatures, as monitored by circular dichroism and fluorescence spectroscopy. It has a highly stable secondary structure over a broad range of pH, undergoes subtle tertiary structure changes at low pH and is less stable under moderate ionic strength conditions. These results bring new insights into the structural properties of OPBTc, contributing to future studies on the rational design of OPBTc inhibitors as a promising strategy for Chagas disease chemotherapy.
authors.affiliation: Pathogen-Host Interface Laboratory, Department of Cell Biology, The University of Brasília, Brasília, Brazil.
authors.forename: Flávia Nader
authors.initials: FN
authors.lastname: Motta
authors.forename: Izabela M D
authors.initials: IM
authors.lastname: Bastos
authors.forename: Eric
authors.initials: E
authors.lastname: Faudry
authors.forename: Christine
authors.initials: C
authors.lastname: Ebel
authors.forename: Meire M
authors.initials: MM
authors.lastname: Lima
authors.forename: David
authors.initials: D
authors.lastname: Neves
authors.forename: Michel
authors.initials: M
authors.lastname: Ragno
authors.forename: João Alexandre R G
authors.initials: JA
authors.lastname: Barbosa
authors.forename: Sônia Maria
authors.initials: SM
authors.lastname: de Freitas
authors.forename: Jaime Martins
authors.initials: JM
authors.lastname: Santana
mesh_terms: D002850:Chromatography, Gel; D002942:Circular Dichroism; D055503:Protein Multimerization; D015800:Protozoan Proteins / Q000737:chemistry* / Q000378:metabolism*; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013696:Temperature; D014349:Trypanosoma cruzi / Q000378:metabolism*; D037521:Virulence Factors / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D015800:Protozoan Proteins; D037521:Virulence Factors; D012697:Serine Endopeptidases; C090703:oligopeptidase B