Isotope-Coded Affinity Tags with Tunable Reactivities for Protein Footprinting

摘要

Fundamental to our understanding of biological systems is the ability to define interaction surfaces and conformational changes in dynamic protein complexes, yet this objective remains a challenge. Protein footprinting is a powerful solution to this problem. Methods such as hydrogen-deuterium exchange, limited proteolysis, and radiolytic cleavage assess changes in protein surface exposure by measuring the susceptibility of the polypeptide backbone to modification. A complementary approach is to use side-chain modification. Because of their unique nucleophilicity, cysteine (Cys) residues at select positions can report on solvent accessibility and local chemical environment. The reactivity of a Cys residue can be measured to define interaction surfaces at individual amino acid resolution. Moreover, methods have been developed for rapid generation of a library of Cys variants for a protein of interest.