A SIMPLE AND SENSITIVE EXPERIMENT FOR MEASUREMENT OF JCC COUPLINGS BETWEEN BACKBONE CARBONYL AND METHYL CARBONS IN ISOTOPICALLY ENRICHED PROTEINS

A simple 2D difference experiment is described that allows quantitativ e measurement of C-13-C-13 J couplings between backbone carbonyl and s ide-chain carbons. Precise 3J(CC) values were measured from data recor ded in just 2 h for a 1-mM solution of the 20-kD complex between the p rotein calmodulin and a 26-residue synthetic peptide. The J couplings aid in determining the chi1 angles of valine, isoleucine and threonine residues, and in making stereospecific assignments of the Val C(gamma ) methyl groups. Error analysis indicates that the uncertainty in the derived J couplings is generally less than ca. 0.3 Hz.