Conformation of a Troponin-I Peptide Bound to Troponin-C as Determined by 1H NMR
The Ca+2-mediated interaction between troponin-l (Tn-I) and troponin-C (Tn-C) in the thin filament of skeletal muscle is an important element in the regulation of the actomyosin ATPase. It has been proposed that the structural changes produced by Ca+2 binding result in the presentation of regions on the surface of Tn-C capable of binding Tn-I with high affinity (1,2,3). In particular, it appears that residues 84–1151 on Tn-C (which represent the N-terminal side of Ca+2-binding site III) interact with residues 97–117 of Tn-I (3,5). A synthetic analog of the inhibitory region of Tn-I, Nα-acetyl-Tn-l(104–115)amide, has been made by solid phase synthesis. This region represents the minimum sequence necessary for inhibition of actomyosin ATPase activity and is found to bind to Tn-C in a Ca+2-dependent fashion, with a binding constant of 1 x 105M-1 (6,7). The following body of work primarily addresses the question of secondary structure adopted by the Tn-I peptide when bound to Tn-C. In order to probe the structure of the bound Tn-I peptide, high field two-dimensional Nuclear Magnetic Resonance (NMR) techniques are used. Possible binding sites on the Tn-C molecule are also considered in light of recent findings from cross-linking studies between rabbit s-Tn-C, to which a maleimide cross-linker molecule has been attached at Cys98, and residues on whole Tn-I (8).
KeywordsNuclear MagnetiC Resonance Distance Restraint Actomyosin ATPase Interproton Distance Ring Pucker
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- 3.P.J. Cachia, J. Gariepy and R.S. Hodges, in: “Calmodulin Antagonists and Cellular Physiology,” Hidaka, H., and Hartshorne, D.J., eds., pp 63–88, Acadmic Press, New York (1985)Google Scholar
- 5.S.V. Perry, “The Regulation of Contractile Activity in Muscle,” Bioc. Soc. Trans. 7: 573 (1977)Google Scholar
- 9.R.R. Ernst, G. Bodenhausen and A. Wokaun, “Principles of Nuclear Magnetic Resonance in One and Two Dimensions,” Clarendon Press, Oxford (1987)Google Scholar
- 10.K. Wüthrich, “NMR of Proteins and Nucleic Acids,” Wiley, New York (1986)Google Scholar
- 13.K. Golosinska, M.J. Carpenter and L.B. Smillie, personal communicationsGoogle Scholar
- 15.B.W. Erickson and R.B. Merrifield, in: “The Proteins,” Neurath, H. and Hill, R.H., eds., Vol II, pp225–527, Academic Press, New York (1976)Google Scholar
- 22.W.F. van Gunsteren and H.J.C. Berendsen, Laboratory of Physical Chemistry, University of Groningen, Nijenborgh16, 9747 Ag Groningen, The NetherlandsGoogle Scholar