Conformational modulation of troponin T by configuration of the NH2-terminal variable region and functional effects.
Troponin T (TnT) is an essential element in the thin filament-based regulatory system of striated muscle. Alternative mRNA splicing generates multiple TnT isoforms with primary structural differences in the NH2-terminal region. The functional significance of this hypervariable NH2-terminal domain and the developmental or muscle type-specific TnT isoforms is not fully understood. We have analyzed chicken breast muscle TnT containing a metal-binding cluster [H(E/A)EAH]4-7 (Tx) in the NH2-terminal region to demonstrate potential effects of the NH2-terminal structure on the conformation of TnT [Ogut, O., and Jin, J.-P. (1996) Biochemistry 35, 16581-16590]. Using specific antibody epitope analysis on this metal-binding TnT model, this study revealed that the binding of Zn2+ to the NH2-terminal region of chicken breast muscle TnT induces extensive conformational changes in the whole protein as demonstrated by a significant decrease in binding avidity of a polyclonal anti-TnT serum which recognizes multiple epitopes on the TnT molecule. This NH2-terminal configuration-based effect is not restricted to the metal ion interaction, whereas the binding of anti-NH2 terminus monoclonal antibodies to TnT induced similar changes. Protein-binding assays have shown that the NH2-terminal variability-induced conformational changes can alter TnT's binding affinity for tropomyosin and troponin I. The results suggest a functional modulation of TnT through the configuration of the NH2-terminal domain, and this novel mechanism may mediate the physiological significance of the TnT isoform regulation.
Department of Physiology and Biophysics, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106-4970, USA.
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D000495:Allosteric Site / Q000276:immunology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001666:Binding Sites, Antibody; D015153:Blotting, Western; D002645:Chickens; D000939:Epitopes / Q000276:immunology; D007106:Immune Sera / Q000378:metabolism; D007536:Isomerism; D008297:Male; D008321:Mammary Glands, Animal; D018482:Muscle, Skeletal; D010446:Peptide Fragments / Q000737:chemistry* / Q000276:immunology / Q000502:physiology; D011485:Protein Binding / Q000276:immunology; D011487:Protein Conformation; D011817:Rabbits; D014335:Tropomyosin / Q000302:isolation & purification / Q000502:physiology; D019210:Troponin I / Q000302:isolation & purification / Q000502:physiology; D020107:Troponin T / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification / Q000502:physiology; D015032:Zinc / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000939:Epitopes; D007106:Immune Sera; D010446:Peptide Fragments; D014335:Tropomyosin; D019210:Troponin I; D020107:Troponin T; D015032:Zinc