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.
Wang
J
J
Department of Physiology and Biophysics, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106-4970, USA.
Jin
J P
JP
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