Using ESI-MS to probe protein structure by site-specific noncovalent attachment of 18-crown-6

ANYL 224

Ryan R Julian, ryan.julian@ucr.edu and Tony Ly, tly003@ucr.edu. Department of Chemistry, University of California, Riverside, Riverside, CA 92521
A new method for probing changes in the equilibrium structures and folding states of proteins will be described. The technique probes side chain availability as determined by a specific interaction between lysine and 18-crown-6 ether (18C6). Various intramolecular interactions which are determined by protein structure are competitive with the lysine/18C6 interaction and can prevent noncovalent attachment of 18C6. Structural rearrangement typically leads to enhanced side chain availability. Experiments conducted with cytochrome c, ubiquitin, and melittin reveal that the method is sensitive to subtle structural changes, including modification of secondary structure. Experiments can be performed under a variety of conditions, including modifications of the pH and amount of organic co-solvent.