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Toxins 2019, 11(4), 194; https://doi.org/10.3390/toxins11040194

Purification and Biochemical Characterization of TsMS 3 and TsMS 4: Neuropeptide-Degrading Metallopeptidases in the Tityus serrulatus Venom

1
Immunochemistry Laboratory, Butantan Institute, São Paulo SP 05503-900, Brazil
2
Biochemistry and Biophysics Laboratory, Butantan Institute, São Paulo SP 05503-900, Brazil
*
Authors to whom correspondence should be addressed.
Received: 1 March 2019 / Revised: 22 March 2019 / Accepted: 28 March 2019 / Published: 31 March 2019
(This article belongs to the Section Animal Venoms)
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Abstract

Although omics studies have indicated presence of proteases on the Tityus serrulatus venom (TsV), little is known about the function of these molecules. The TsV contains metalloproteases that cleave a series of human neuropeptides, including the dynorphin A (1-13) and the members of neuropeptide Y family. Aiming to isolate the proteases responsible for this activity, the metalloserrulase 3 and 4 (TsMS 3 and TsMS 4) were purified after two chromatographic steps and identified by mass spectrometry analysis. The biochemical parameters (pH, temperature and cation effects) were determined for both proteases, and the catalytic parameters (Km, kcat, cleavage sites) of TsMS 4 over fluorescent substrate were obtained. The metalloserrulases have a high preference for cleaving neuropeptides but presented different primary specificities. For example, the Leu-enkephalin released from dynorphin A (1-13) hydrolysis was exclusively performed by TsMS 3. Neutralization assays using Butantan Institute antivenoms show that both metalloserrulases were well blocked. Although TsMS 3 and TsMS 4 were previously described through cDNA library studies using the venom gland, this is the first time that both these toxins were purified. Thus, this study represents a step further in understanding the mechanism of scorpion venom metalloproteases, which may act as possible neuropeptidases in the envenomation process. View Full-Text
Keywords: Tityus serrulatus; metalloserrulases; proteases; purification; biochemical characterization; neuropeptides Tityus serrulatus; metalloserrulases; proteases; purification; biochemical characterization; neuropeptides
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited (CC BY 4.0).

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Cajado-Carvalho, D.; da Silva, C.C.F.; Kodama, R.T.; Mariano, D.O.C.; Pimenta, D.C.; Duzzi, B.; Kuniyoshi, A.K.; Portaro, F.V. Purification and Biochemical Characterization of TsMS 3 and TsMS 4: Neuropeptide-Degrading Metallopeptidases in the Tityus serrulatus Venom. Toxins 2019, 11, 194.

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