title: Measurement of the exchange rates of rapidly exchanging amide protons: application to the study of calmodulin and its complex with a myosin light chain kinase fragment.
abstract: A technique is described for measuring the approximate exchange rates of the more labile amide protons in a protein. The technique relies on a comparison of the intensities in 1H-15N correlation spectra recorded with and without presaturation of the water resonance. To distinguish resonance attenuation caused by hydrogen exchange from attenuation caused by cross relaxation, the experiment is repeated at several different pH values and the difference in attenuation of any particular amide resonance upon presaturation is used for calculating its exchange rate. The technique is demonstrated for calmodulin and for calmodulin complexed with its binding domain of skeletal muscle myosin light chain kinase. Upon complexation, increased amide exchange rates are observed for residues Lys75 through Thr79 located in the 'central helix' of calmodulin, and for the C-terminal residues Ser147 and Lys148. In contrast, a decrease in amide exchange rate is observed at the C-terminal end of the F helix, from residues Thr110 through Glu114.
authors.affiliation: Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892.
authors.forename: S
authors.initials: S
authors.lastname: Spera
authors.forename: M
authors.initials: M
authors.lastname: Ikura
authors.forename: A
authors.initials: A
authors.lastname: Bax
mesh_terms: D000577:Amides / Q000378:metabolism; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism; D000818:Animals; D002147:Calmodulin / Q000378:metabolism*; D003903:Deuterium / Q000378:metabolism; D006859:Hydrogen / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D011485:Protein Binding; D011522:Protons; D011817:Rabbits
chemical_list: D000577:Amides; D000596:Amino Acids; D002147:Calmodulin; D010446:Peptide Fragments; D011522:Protons; D006859:Hydrogen; D003903:Deuterium; D009219:Myosin-Light-Chain Kinase