Ferraz Karla Fernanda, De Lucca Caetano Lhiri Hanna, Orefice Daniele Pereira, Calabria Paula Andreia Lucas, Della-Casa Maisa Splendore, Freitas-de-Sousa Luciana Aparecida, Beraldo-Neto Emidio, Sanabani Sabri Saeed, Magalhães Geraldo Santana, Clissa Patricia Bianca
Immunopathology Laboratory, Butantan Institute, São Paulo 05585-090, Brazil.
Biochemistry Laboratory, Butantan Institute, São Paulo 05503-900, Brazil.
Toxins (Basel). 2024 Dec 3;16(12):524. doi: 10.3390/toxins16120524.
Jararhagin-C (JarC) is a protein from the venom of consisting of disintegrin-like and cysteine-rich domains. JarC shows a modulating effect on angiogenesis and remodeling of extracellular matrix constituents, improving wound healing in a mouse experimental model. JarC is purified from crude venom, and the yield is less than 1%. The aim of this work was to obtain the recombinant form of JarC and to test its biological activity. For this purpose, the bicistronic vector pSUMOUlp1 was used. This vector allowed the expression of the recombinant toxin JarC (rJarC) in fusion with the small ubiquitin-related modifier (SUMO) as well as the SUMO protease Ulp1. After expression, this protease was able to efficiently remove SUMO from rJarC inside the bacteria. rJarC free from SUMO was purified at the expected molecular mass and recognized by polyclonal anti-jararhagin antibodies. In terms of biological activity, both the native and recombinant forms showed no toxicity to the HUVEC cell line CRL1730 and were effective in modulating cell migration activity in the experimental in vitro model. These results demonstrate the successful production of rJarC and the preservation of its biological activity, which may facilitate further investigations into the therapeutic potential of this snake venom-derived protein.
矛头蝮蛇毒素C（JarC）是一种来自矛头蝮蛇毒液的蛋白质，由类去整合素结构域和富含半胱氨酸的结构域组成。JarC对血管生成和细胞外基质成分的重塑具有调节作用，在小鼠实验模型中可促进伤口愈合。JarC是从粗毒液中纯化得到的，产量不到1%。这项工作的目的是获得JarC的重组形式并测试其生物活性。为此，使用了双顺反子载体pSUMOUlp1。该载体能够使重组毒素JarC（rJarC）与小泛素相关修饰物（SUMO）以及SUMO蛋白酶Ulp1融合表达。表达后，这种蛋白酶能够在细菌内有效地从rJarC上去除SUMO。去除SUMO的rJarC在预期分子量下得到纯化，并能被多克隆抗矛头蝮蛇毒素抗体识别。在生物活性方面，天然形式和重组形式对人脐静脉内皮细胞系CRL1730均无毒性，并且在体外实验模型中均能有效调节细胞迁移活性。这些结果证明了rJarC的成功制备及其生物活性的保留，这可能有助于进一步研究这种蛇毒衍生蛋白的治疗潜力。
