Otimização do processo de obtenção de uma desintegrina RGD recombinante (DISBA-01) inibidora da integrina ∝vβ3

Abstract

Heterologous expression of recombinant proteins in Escherichia coli generally occurs under denaturing conditions, which can prevent natural protein folding and favor the formation of inclusion bodies, requiring the use of agents such as urea during purification. However, this process ultimately leads to the need to remove the urea at the end, which can result in a loss in the recovery of the biological activity of the protein of interest. Therefore, this work aimed to optimize the production of the recombinant protein DisBa-01, which belongs to the class of disintegrins and exhibits high affinity for the αvβ3 integrin due to its RGD domain, using non-denaturing conditions. With this approach, the idea was to adapt expression and purification steps to maintain the protein in its native form, seeking strategies to reduce the temperature and increase the expression time of microbial culture, avoiding the use of high amounts of urea and thus reducing losses throughout the process and decreasing the cost of its production. The results obtained were promising, as the recombinant protein DisBa-01 was obtained in soluble form, and its biological activity was significant, confirmed by in vitro assays. Overall, the study presents an alternative for the more efficient production of DisBa-01, aiming to facilitate future functional analyses of this disintegrin.