BackgroundThe neglected human diseases caused by trypanosomatids are currently treated with toxic therapy with limited efficacy. In search for novel anti-trypanosomatid agents, we showed previously that the Crotalus viridis viridis (Cvv) snake venom was active against infective forms of Trypanosoma cruzi. Here, we describe the purification of crovirin, a cysteine-rich secretory protein (CRISP) from Cvv venom with promising activity against trypanosomes and Leishmania.Methodology/Principal FindingsCrude venom extract was loaded onto a reverse phase analytical (C8) column using a high performance liquid chromatographer. A linear gradient of water/acetonitrile with 0.1% trifluoroacetic acid was used. The peak containing the isolated protein (confirmed by SDS-PAGE and mass spectrometry) was collected and its protein content was measured. T. cruzi trypomastigotes and amastigotes, L. amazonensis promastigotes and amastigotes and T. brucei rhodesiense procyclic and bloodstream trypomastigotes were challenged with crovirin, whose toxicity was tested against LLC-MK2 cells, peritoneal macrophages and isolated murine extensor digitorum longus muscle. We purified a single protein from Cvv venom corresponding, according to Nano-LC MS/MS sequencing, to a CRISP of 24,893.64 Da, henceforth referred to as crovirin. Human infective trypanosomatid forms, including intracellular amastigotes, were sensitive to crovirin, with low IC50 or LD50 values (1.10–2.38 µg/ml). A considerably higher concentration (20 µg/ml) of crovirin was required to elicit only limited toxicity on mammalian cells.ConclusionsThis is the first report of CRISP anti-protozoal activity, and suggests that other members of this family might have potential as drugs or drug leads for the development of novel agents against trypanosomatid-borne neglected diseases.
Productivity of sugarcane (Saccharum spp.) crops varies at each cutting stage, reaching critical rates close to the fifth cut (fourth ratoon). Knowledge of proteins involved in the regrowth of sugarcane within the cutting process is important for the development of cultivars with greater longevity. The present study presents new information that the proteome of axillary buds is changed in successive cuts in sugarcane culture. Proteins were identified by UPLC-ESI-Q-TOF (ultra-high-performance liquid chromatography coupled with electrospray ionisation–quadrupole–time-of-flight) mass spectrometry and the Mascot tool. A reduction in the number of proteins was evident in the axillary buds of the fifth cut, as well as a reduction in the number of proteins exclusively detected in the axillary buds with the first cut, an indicator of reduction in the expression of genes that may be essential for the stability of culture development. The reduction in agricultural productivity, sprouting and tillering at advanced stages of the sugarcane crop is accompanied by alterations in axillary-bud gene expression, where <50% of the proteins (47.65%) were detected in both the first (plant cane) and in the fifth (fourth ratoon) cutting stage, whereas >50% (52.35%) were expressed in either the axillary buds of the plant cane or the axillary buds of the fourth ratoon. All MS data are available via jPOST and ProteomeXchange with identifiers JPST000331 and PXD007957, respectively.
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