Folding and function of the troponin tail domain. Effects of cardiomyopathic troponin T mutations.
Troponin contains a globular Ca(2+)-binding domain and an elongated tail domain composed of the N terminus of subunit troponin T (TnT). The tail domain anchors troponin to tropomyosin and actin, modulates myosin function, and is a site of cardiomyopathy-inducing mutations. Critical interactions between tropomyosin and troponin are proposed to depend on tail domain residues 112-136, which are highly conserved across phyla. Most cardiomyopathy mutations in TnT flank this region. Three such mutations were examined and had contrasting effects on peptide TnT-(1-156), promoting folding and thermal stability assessed by circular dichroism (F110I) or weakening folding and stability (T104V and to a small extent R92Q). Folding of both TnT-(1-156) and whole troponin was promoted by replacing bovine TnT Thr-104 with human TnT Ala-104, further indicating the importance of this cardiomyopathy site residue for protein folding. Mutation F110I markedly stabilized the troponin tail but weakened binding of holo-troponin to actin-tropomyosin 8-fold, suggesting that loss of flexibility impairs troponin tail function. The effect of the F110I mutation on troponin-tropomyosin binding to actin was much less, indicating this flexibility is particularly important for the interactions of troponin with tropomyosin. We suggest that most cardiomyopathic mutations in the troponin tail alter muscle function indirectly, by perturbing interactions between troponin and tropomyosin requisite for the complex effects of these proteins on myosin.
Hinkle
Ashley
A
Department of Internal Medicine, University of Iowa, Iowa City, Iowa 52242, USA.
Tobacman
Larry S
LS
D000595:Amino Acid Sequence; D000818:Animals; D009202:Cardiomyopathies / Q000235:genetics* / Q000378:metabolism; D002417:Cattle; D002942:Circular Dichroism; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D009206:Myocardium / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D011489:Protein Denaturation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013696:Temperature; D014335:Tropomyosin / Q000235:genetics / Q000378:metabolism; D020107:Troponin T / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D011994:Recombinant Proteins; D014335:Tropomyosin; D020107:Troponin T