Contemporary Enzyme-Based Methods for Recombinant Proteins In Vitro Phosphorylation

Phosphorylation is a reversible, enzyme-controlled posttranslational process affecting approximately one-third of all proteins in eukaryotic cells at any given time. Any deviation in the degree and/or site of phosphorylation leads to an abnormal conformation of proteins, resulting in a decline or loss of their function. Knowledge of phosphorylation-related pathways is essential for understanding the understanding of the disease pathogenesis and for the design of new therapeutic strategies. Recent availability of various kinases at an affordable price differs in activity, specificity, and stability and provides the opportunity of studying and modulating this reaction in vitro. We can exploit this knowledge for other applications. There is an enormous potential to produce fully decorated and active recombinant proteins, either for biomedical or cosmetic applications. Closely related is the possibility to exploit current achievements and develop new safe and efficacious vaccines, drugs, and immunomodulators. In this review, we outlined the current enzyme-based possibilities for in vitro phosphorylation of peptides and recombinant proteins and the added value that immobilized kinases provide.

Automated summary

Phosphorylation is a key posttranslational process that affects protein function, and understanding it is crucial for studying disease pathogenesis and designing new therapeutic strategies. Affordable kinases with varying activity, specificity, and stability offer opportunities for in vitro research and modulation of phosphorylation reactions, facilitating the production of active recombinant proteins.