title: Rhipicephalus sanguineus trypsin inhibitors present in the tick larvae: isolation, characterization, and partial primary structure determination.
abstract: Blood sucking animals are a rich source of proteinase inhibitors mainly those that interfere in their host hemostatic systems. The tick Rhipicephalus sanguineus is an ectoparasite of dogs and other animals. The aims of this work were the purification and characterization of serine proteinase inhibitors present in R. sanguineus larvae (RsTI). The inhibitors (RsTI) were isolated by affinity chromatography on trypsin-Sepharose and ion exchange chromatographies in Resource Q and Mono S columns. These RsTIs were separated in around 12 different protein peaks, when they showed molecular masses between 8 and 18 kDa, by SDS-PAGE. Purified RsTIs presented differences in the specificity for different serine proteinases. RsTIQ2 was, better inhibitor than RsTIQ7 and RsTIS5 for neutrophil elastase, plasmin, and HuPK with dissociation constants (K(i)) of 1.3, 3.2, and 22 nM, respectively. Other inhibitors such as RsTIQ7, RsTIS3, and RsTIS5 also affected neutrophil elastase and plasmin with K(i) in the nM range. The RsTIQ2, RsTIQ7, and RsTIS5 amino acid sequence data allowed classifying them as members of the Kunitz-type serine proteinase inhibitor family, even though the RsTI role is still unknown. Our present results showed that serine proteinase inhibitors from R. sanguineus are similar to inhibitors from Boophilus microplus other hard tick species, suggesting a similar role of these inhibitors in hard tick species and also as a potential tool to generate or improve vaccine against different ectoparasites with an unique antigen.
authors.affiliation: Departamento de Bioquímica, Universidade Federal de São Paulo, Escola Paulista de Medicina, 04044-020 São Paulo, SP, Brazil.
authors.forename: Simone
authors.initials: S
authors.lastname: Sant'Anna Azzolini
authors.forename: Sergio Daishi
authors.initials: SD
authors.lastname: Sasaki
authors.forename: Ricardo José Soares
authors.initials: RJ
authors.lastname: Torquato
authors.forename: Renato
authors.initials: R
authors.lastname: Andreotti
authors.forename: Eliane
authors.initials: E
authors.lastname: Andreotti
authors.forename: Aparecida Sadae
authors.initials: AS
authors.lastname: Tanaka
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D004789:Enzyme Activation; D026863:Ixodidae / Q000737:chemistry* / Q000378:metabolism; D007814:Larva / Q000737:chemistry; D008969:Molecular Sequence Data; D011487:Protein Conformation; D015842:Serine Proteinase Inhibitors; D014361:Trypsin Inhibitors / Q000737:chemistry* / Q000145:classification / Q000302:isolation & purification* / Q000378:metabolism
chemical_list: D015842:Serine Proteinase Inhibitors; D014361:Trypsin Inhibitors