title: Comparative studies on the functional roles of N- and C-terminal regions of molluskan and vertebrate troponin-I.
abstract: Vertebrate troponin regulates muscle contraction through alternative binding of the C-terminal region of the inhibitory subunit, troponin-I (TnI), to actin or troponin-C (TnC) in a Ca(2+)-dependent manner. To elucidate the molecular mechanisms of this regulation by molluskan troponin, we compared the functional properties of the recombinant fragments of Akazara scallop TnI and rabbit fast skeletal TnI. The C-terminal fragment of Akazara scallop TnI (ATnI(232-292)), which contains the inhibitory region (residues 104-115 of rabbit TnI) and the regulatory TnC-binding site (residues 116-131), bound actin-tropomyosin and inhibited actomyosin-tropomyosin Mg-ATPase. However, it did not interact with TnC, even in the presence of Ca(2+). These results indicated that the mechanism involved in the alternative binding of this region was not observed in molluskan troponin. On the other hand, ATnI(130-252), which contains the structural TnC-binding site (residues 1-30 of rabbit TnI) and the inhibitory region, bound strongly to both actin and TnC. Moreover, the ternary complex consisting of this fragment, troponin-T, and TnC activated the ATPase in a Ca(2+)-dependent manner almost as effectively as intact Akazara scallop troponin. Therefore, Akazara scallop troponin regulates the contraction through the activating mechanisms that involve the region spanning from the structural TnC-binding site to the inhibitory region of TnI. Together with the observation that corresponding rabbit TnI-fragment (RTnI(1-116)) shows similar activating effects, these findings suggest the importance of the TnI N-terminal region not only for maintaining the structural integrity of troponin complex but also for Ca(2+)-dependent activation.
authors.affiliation: Laboratory of Biotechnology and Microbiology, Graduate School of Fisheries Sciences, Hokkaido University, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Tanaka
authors.forename: Yuhei
authors.initials: Y
authors.lastname: Takeya
authors.forename: Teppei
authors.initials: T
authors.lastname: Doi
authors.forename: Fumiaki
authors.initials: F
authors.lastname: Yumoto
authors.forename: Masaru
authors.initials: M
authors.lastname: Tanokura
authors.forename: Iwao
authors.initials: I
authors.lastname: Ohtsuki
authors.forename: Kiyoyoshi
authors.initials: K
authors.lastname: Nishita
authors.forename: Takao
authors.initials: T
authors.lastname: Ojima
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D017301:Ca(2+) Mg(2+)-ATPase / Q000037:antagonists & inhibitors; D002118:Calcium / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics; D066298:In Vitro Techniques; D007700:Kinetics; D008969:Molecular Sequence Data; D008974:Mollusca; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011817:Rabbits; D013045:Species Specificity; D014335:Tropomyosin / Q000378:metabolism; D019209:Troponin C / Q000378:metabolism; D019210:Troponin I / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D000199:Actins; D018076:DNA, Complementary; D010446:Peptide Fragments; D014335:Tropomyosin; D019209:Troponin C; D019210:Troponin I; D017301:Ca(2+) Mg(2+)-ATPase; D002118:Calcium