Recombinant protein expression: Challenges in production and folding related matters

Protein folding is a biophysical process by which proteins reach a specific three-dimensional structure. The amino acid sequence of a polypeptide chain contains all the information needed to determine the final three-dimensional structure of a protein. When producing a recombinant protein, several problems can occur, including proteolysis, incorrect folding, formation of inclusion bodies, or protein aggregation, whereby the protein loses its natural structure. To overcome such limitations, several strategies have been developed to address each specific issue. Identification of proper protein refolding conditions can be challenging, and to tackle this high throughput screening for different recombinant protein folding conditions can prove a sound solution. Different approaches have emerged to tackle refolding issues. One particular approach to address folding issues involves molecular chaperones, highly conserved proteins that contribute to proper folding by shielding folding proteins from other proteins that could hinder the process. Proper protein folding is one of the main prerequisites for post-translational modifications. Incorrect folding, if not dealt with, can lead to a buildup of protein misfoldings that damage cells and cause widespread abnormalities. Said post-translational modifications, widespread in eukaryotes, are critical for protein structure, function and biological activity. Incorrect post-translational protein modifications may lead to individual consequences or aggregation of therapeutic proteins. In this review article, we have tried to examine some key aspects of recombinant protein expression. Accordingly, the relevance of these proteins is highlighted, major problems related to the production of recombinant protein and to refolding issues are pinpointed and suggested solutions are presented. An overview of post-translational modification, their biological significance and methods of identification are also provided. Overall, the work is expected to illustrate challenges in recombinant protein expression.