The peptide pattern obtained during degradation of TN-C component of troponin by trypsin markedly depends on the concentration of calcium ions. In the presence of calcium higher than 0.1 mM a peptide ...with a mass of about 16 000 daltons is formed, followed by its splitting into two peptides (8500 and 7500 daltons), which accumulate during further digestion. When calcium is sequestred by EGTA the degradation of TN-C is much faster. The first products of digestion are the 11 000 and 7500 dalton peptides. The former one is degraded to 9700 dalton peptide and, subsequently, both peptides are split to small fragments. In the presence of calcium only three -NH2 groups per molecule of TN-C appear during digestion, whereas in its absence almost all peptide bonds available for trypsin are cleaved. The fragments of TN-C obtained in the presence of calcium are still able to interact with TN-I component as judged by disc-electrophoresis performed in the presence of calcium. During digestion in the presence of calcium only a slight decrease of tyrosyl intrinsic fluorescence occurs and the further decrease after removal of calcium bound to TN-C is fully reversible. Similarly, the content of alpha-helix decreases only slowly during digestion in the presence of calcium. All the results suggest that calcium stabilizes the structure of TN-C molecule, so that trypsin in the presence of calcium releases one small peptide from N-terminal not involved in the first "EF-hand" and the other from the region between pairs 1-2 and 3-4 of "EF-hands" in the nomenclature of Kretsinger (see Kretsinger and Barry 16). The obtained fragments, containing almost exclusively pairs of "EF-hands", restore most of the properties of original TN-C molecule.
Proton magnetic resonance spectroscopy has been used to study the cation(Mg2+, Ca2+)‐dependent conformational states of the C‐terminal domain of rabbit skeletal troponin C under a variety of solution ...conditions. Nuclear Overhauser data and paramagnetic probe observations provide definition of the configuration of this region of troponin C. Comparative study of homologous proteins identify common features of the tertiary structure relevant to the cation binding reaction. Complex formation with troponin I and the drug trifuloperazine is observed to adjust the solution conformation of the C‐terminal domain of troponin C. The interactive conformational response to cation coordination and of the binding of the drug and troponin I are discussed.
Proton magnetic resonance spectroscopy has been used to study the cation (Mg2+, Ca2+)-dependent conformational states of the C-terminal domain of rabbit skeletal troponin C under a variety of ...solution conditions. Nuclear Overhauser data and paramagnetic probe observations provide definition of the configuration of this region of troponin C. Comparative study of homologous proteins identify common features of the tertiary structure relevant to the cation binding reaction. Complex formation with troponin I and the drug trifluoperazine is observed to adjust the solution conformation of the C-terminal domain of troponin C. The interactive conformational response to cation coordination and the binding of the drug and troponin I are discussed.
The relaxation rate enhancements of the 23Na nuclei for NaHCO3 solutions of troponin C and its tryptic peptides TR‐1 and TR‐2 indicate true binding of Na+ ions to these biomolecules. The low‐affinity ...sites I and II of TR‐1 and troponin C are the sites of competitive Na+/Ca2+ binding, below one calcium ion per molecule, with log KNa∼ 2. At low calcium content Na+ ions bind to TR‐2 and to troponin C non‐competitively with Ca2+ ions; binding of Ca2+ ions to the high‐affinity sites III and IV allosterically affects the binding of the Na+ ions: even when sites I and II, located on TR‐1 or sites I, II. III, IV of troponin C, are saturated with Ca2+ ions, Na+ ions continue to bind weakly at secondary binding sites.
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