Biotecnological applications of plastid thioredoxins in molecular pharming

Abstract

Thioredoxins (Trxs) are small ubiquitous disulfide reductases !ha! play importan! roles in the redox regu!ation of many cellular processes, even though sorne redox-independent functions, such as chaperone activity, have also been attributed to them. From a biotechnological point of view, Trxs are widely known to enhance expression and solubility of recombinant proteins in microbial expression systems. In this context, and given the common ancestor of chloroplasts and bacteria, we have explored whether plastid Trxs could actas modulators of recombinant protein expression in transgenic chloroplasts. For !ha! purpose, two tobacco Trxs (m and f) with different phylogenetic origins were assessed. Using plastid transformation, we assayed both fusion and co-expression ofTrxs with human serum albumin (HSA) and human cardiotrophin-1 (CT -1 ), two proteins with high therapeutic potential. Our results show that !he fusion approach markedly in creases the final yield of both recombinant proteins, likely due to a high stability of !he fusion protein. Co-expression of HSA with Trxs prevented the formation of large protein bodies within chloroplasts. Additionally, co-expression of CT-1 with Trxs improved the bioactivity of the recombinant CT-1 produced in tobacco leaves. Given that we have demonstrated the in vitro chaperone activity of Trx m and f, we propase a direct relationship between Trx overexpression and HSA aggregates solubilization or CT-1 functionality improvement into the chloroplast.