Disease-causing mutations in cardiac troponin T: Identification of a critical tropomyosin-binding region

Fifteen percent of the mutations causing familial hypertrophic cardiomyopat hy are in the troponin T gene. Most mutations are clustered between residue s 79 and 179, a region known to bind to tropomyosin at the C-terminus near the complex between the N- and C-termini. Nine mutations were introduced in to a troponin T fragment, Gly-hcTnT(70-170), that is soluble, alpha -helica l, binds to tropomyosin, promotes the binding of tropomyosin to actin, and stabilizes an overlap complex of N-terminal and C-terminal tropomyosin pept ides. Mutations between residues 92 and 110 (Arg92Leu, Arg92Gln, Arg92Trp, Arg94Leu, Ala104Val, and Phe110lle) impair tropomyosin-dependent functions of troponin T. Except for Ala104Val, these mutants bound less strongly to a tropomyosin affinity column and were less able to stabilize the TM overlap complex, effects that were correlated with increased stability of the trop onin T, measured using circular dichroism. All were less effective in promo ting the binding of tropomyosin to actin. Mutations within residues 92-110 may cause disease because of altered interaction with tropomyosin at the ov erlap region, critical for cooperative actin binding and regulatory functio n. A model for a five-chained coiled-coil for troponin T in the tropomyosin overlap complex is presented. Mutations outside the region (lle79Asn, Delt a 160Glu, and Glu163Lys) functioned normally and must cause disease by anot her mechanism.