title: Structure and interactions of the carboxyl terminus of striated muscle alpha-tropomyosin: it is important to be flexible.
abstract: Tropomyosin (TM) binds to and regulates the actin filament. We used circular dichroism and heteronuclear NMR to investigate the secondary structure and interactions of the C terminus of striated muscle alpha-TM, a major functional determinant, using a model peptide, TM9a(251-284). The (1)H(alpha) and (13)C(alpha) chemical shift displacements show that residues 252 to 277 are alpha-helical but residues 278 to 284 are nonhelical and mobile. The (1)H(N) and (13)C' displacements suggest that residues 257 to 269 form a coiled coil. Formation of an "overlap" binary complex with a 33-residue N-terminal chimeric peptide containing residues 1 to 14 of alpha-TM perturbs the (1)H(N) and (15)N resonances of residues 274 to 284. Addition of a fragment of troponin T, TnT(70-170), to the binary complex perturbs most of the (1)H(N)-(15)N cross-peaks. In addition, there are many new cross-peaks, showing that the binding is asymmetric. Q263, in a proposed troponin T binding site, shows two sets of side-chain (15)N-(1)H cross-peaks, indicating conformational flexibility. The conformational equilibrium of the side chain changes upon formation of the binary and ternary complexes. Replacing Q263 with leucine greatly increases the stability of TM9a(251-284) and reduces its ability to form the binary and ternary complexes, showing that conformational flexibility is crucial for the binding functions of the C terminus.
authors.affiliation: University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, 675 Hoes Lane, Piscataway, NJ 08854-5635, USA. greenfie@rwja.umdnj.edu
authors.forename: Norma J
authors.initials: NJ
authors.lastname: Greenfield
authors.forename: Thomas
authors.initials: T
authors.lastname: Palm
authors.forename: Sarah E
authors.initials: SE
authors.lastname: Hitchcock-DeGregori
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D055592:Biophysical Phenomena; D001703:Biophysics; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D010452:Peptide Biosynthesis; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000378:metabolism; D013696:Temperature; D014335:Tropomyosin / Q000737:chemistry*
chemical_list: D010455:Peptides; D011994:Recombinant Proteins; D014335:Tropomyosin