Developmentally Regulated Protein Expression Mediated by Dimethylsulfoxide in Herpetomonas samuelpessoai

Abstract

We have analyzed the total cell extract, cell surface, and secretory protein profiles related to cellular differentiation triggered by dimethylsulfoxide in the insect trypanosomatid Herpetomonas samuelpessoai. The flagellates were cultivated in chemically defined conditions in the absence or in the presence of 4% DMSO, and the resolved protein bands were detected by SDS-PAGE gels and avidin-Western blotting. The cell-associated proteins showed a complex pattern of around 40 silver-staining bands ranging from 15 to 150 kDa. There were generally minor quantitative differences in the protein profile between the non-treated and the DMSO-treated cells. The cell-surface protein profile revealed by the incubation of live parasites with biotin showed a decrease in the expression of the 120 kDa biotinylated polypeptide observed in the DMSO-treated cells when compared with untreated ones. However, control samples of both systems showed an endogenous biotinylated polypeptide of 63 kDa which also presented gelatinolytic activity. The trypanosomatids released at least 10 polypeptides to the culture medium. A low molecular mass exopolypeptide (35 kDa) was found exclusively in untreated cells, whereas a high-molecular-mass exopolypetide (250 kDa) was preferentially found in DMSO-treated cells.

The ubiquitous gp63-like metalloprotease from lower trypanosomatids: in the search for a function

“…Differences in migration of the gp63 on SDS-PAGE could be explained by a heterogeneity resulting from posttranslational modifi cations of a common precursor, such as glycosylation (Bordier et al 1986). In this context, the gp63 protein obtained from H. samuelpessoai was able to bind to Concanavalin A agglutinin; this fact can be attributed to its glycoprotein nature (Santos et al 2001b). Curiously, H. samuelpessoai gp63 is also endogenously biotinylated (Santos et al 2001b).…”

“…In this context, the gp63 protein obtained from H. samuelpessoai was able to bind to Concanavalin A agglutinin; this fact can be attributed to its glycoprotein nature (Santos et al 2001b). Curiously, H. samuelpessoai gp63 is also endogenously biotinylated (Santos et al 2001b).…”

Biological Roles of Peptidases in Trypanosomatids~!2009-11-26~!2010-02-15~!2010-03-18~!

“…It is well known that the symbiont interferes with several aspects of the host trypanosomatid physiology [233], including the production of proteolytic enzymes [10,23,26]. In this sense, previous studies showed that endosymbiont-bearing strains are more efficient in binding to insect cell lines and midguts, as compared to symbiotic free strains, which is probably related to differential expression of surface molecules [27,234].…”

Applications of Zymography (Substrate-SDS-PAGE) for Peptidase Screening in a Post-Genomic Era

“…The 66 kDa metallopeptidase activity was detected in the parasite membrane fraction after Triton X-114 partition (Etges, 1992;Schneider and Glaser, 1993;Santos et al 2003) or after treatment of living cells with phospholipase C (Santos et al 2002;Santos et al 2006) as well as in the extracellular environment as the major secreted peptidase component (Santos et al 2001(Santos et al , 2003Elias et al 2006). This metallopeptidase produced by H. samuelpessoai cells shares common biochemical and immunological properties (Elias et al 2006;Santos et al 2006) with the major metallopeptidase expressed by Leishmania species, called leishmanolysin or gp63, a virulence factor that participates in different stages of the parasite life cycle such as adhesion and escape from host immune response (Yao, 2010).…”