STRUCTURE AND INTERACTION SITE OF THE REGULATORY DOMAIN OF TROPONIN-CWHEN COMPLEXED WITH THE 96-148 REGION OF TROPONIN-I

The structure of the regulatory domain of chicken skeletal troponin-C (residues 1-90) when complexed with the major inhibitory region (resid ues 96-148) of chicken skeletal troponin-I was determined using multin uclear, multidimensional NMR spectroscopy. This complex represents the first interaction formed between the regulatory domain of troponin-C and troponin-I after calcium binding in the regulation of muscle contr action. The stoichiometry of the complex was determined to be I:I, wit h a dissociation constant in the 1-40 mu M range. The structure of tro ponin-C in the complex was calculated from 1039 NMR distance and 111 d ihedral angle restraints. When compared to the structure of this domai n in the calcium saturated ''open'' form but in the absence of troponi n-I, the bound structure appears to be slightly more ''closed''. The t roponin-I peptide-binding site was found to be in the hydrophobic pock et of calcium saturated troponin-C, using edited/filtered NMR experime nts and chemical shift mapping of changes induced in the regulatory do main upon peptide binding. The troponin-I peptide (residues 96-148) wa s found to bind to the regulatory domain of troponin-C very similarly, but not identically, to a shorter troponin-I peptide (region 115-131) thought to represent the major interaction site of troponin-I for thi s domain of troponin-C.