Many animal toxins may target the same molecules that need to be controlled in certain pathologies; therefore, some toxins have led to the formulation of drugs that are presently used, and many other drugs are still under development. Nevertheless, collecting sufficient toxins from the original source might be a limiting factor in studying their biological activities. Thus, molecular biology techniques have been applied in order to obtain large amounts of recombinant toxins into Escherichia coli. However, most animal toxins are difficult to express in this system, which results in insoluble, misfolded, or unstable proteins. To solve these issues, toxins have been fused with tags that may improve protein expression, solubility, and stability. Among these tags, the SUMO (small ubiquitin-related modifier) has been shown to be very efficient and can be removed by the Ulp1 protease. However, removing SUMO is a labor- and time-consuming process. To enhance this system, here we show the construction of a bicistronic vector that allows the expression of any protein fused to both the SUMO and Ulp1 protease. In this way, after expression, Ulp1 is able to cleave SUMO and leave the protein interest-free and ready for purification. This strategy was validated through the expression of a new phospholipase D from the spider Loxosceles gaucho and a disintegrin from the Bothrops insularis snake. Both recombinant toxins showed good yield and preserved biological activities, indicating that the bicistronic vector may be a viable method to produce proteins that are difficult to express.

Angiogenesis, the formation of new blood vessels, plays a critical role in various physiological and pathological conditions. Snake venom disintegrins (SVDs) have been identified as significant regulators of this process. In this review, we explore the dual roles of SVD in angiogenesis, both as antiangiogenic agents by inhibiting integrin binding and interfering with vascular endothelial growth factors and as proangiogenic agents by enhancing integrin binding, stimulating cell migration and proliferation, and inducing neoangiogenesis. Studies in vitro and in animal models have demonstrated these effects and offer significant therapeutic opportunities. The potential applications of SVD in diseases related to angiogenesis, such as cancer, ocular diseases, tissue regeneration, wound healing, and cardiovascular diseases, are also discussed. Overall, SVDs are promising potential therapeutics, and further advances in this field could lead to innovative treatments for diseases related to angiogenesis.

Jararhagin-C (JarC) is a protein from the venom of Bothrops jararaca 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.

Jararhagin-C (JarC) is a protein from the venom of Bothrops jararaca 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 mice 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 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.

Angiogenesis is a critical process in various physiological and pathological conditions, and snake venom disintegrins (SVDs) have been shown to be important regulators of this process. In this review, we explore the dual roles of SVD in angiogenesis, both as antiangiogenic agents by inhibiting integrin binding and interfering with vascular endothelial growth factors and as proangiogenic agents by enhancing integrin binding, stimulating cell migration and proliferation, and inducing neoangiogenesis. Studies in vitro and in animal models have demonstrated these effects and offer significant therapeutic opportunities. The potential applications of SVD in diseases related to angiogenesis, such as cancer, ocular diseases, tissue regeneration, wound healing, and cardiovascular diseases, are also discussed. Overall, SVDs are promising potential therapeutics, and further advances in this field could lead to innovative treatments for diseases related to angiogenesis.