title: Charged residue changes in the carboxy-terminus of alpha-tropomyosin alter mouse cardiac muscle contractility.
abstract: Striated muscle tropomyosin (TM) is an essential thin filament protein that is sterically and allosterically involved in calcium-mediated cardiac contraction. We have previously shown that overexpressing the beta-TM isoform in mouse hearts leads to physiological changes in myocardial relaxation and Ca(2+) handling of myofilaments. Two important charge differences in beta-TM compared to alpha-TM are the exchange of serine and histidine at positions 229 and 276 with glutamic acid and asparagine, respectively, imparting a more negative charge to beta-TM relative to alpha-TM. Our hypothesis is that the net charge at specific sites on TM might be a major determinant of its role in modulating cardiac muscle performance and in regulating Ca(2+) sensitivity of the myofilaments. To address this, we generated transgenic (TG) double mutation mouse lines (alpha-TM DM) expressing mutated alpha-TM at the two residues that differ between alpha- and beta-TM (Ser229Glu + His276Asn). Molecular analyses show 60-88% of the native TM is replaced with alpha-TM DM in the different TG lines. Work-performing heart analyses show that alpha-TM DM mouse hearts exhibit decreased rates of pressure development and relaxation (+dP/dt and -dP/dt). Skinned myofibre preparations from the TG hearts indicate a decrease in calcium sensitivity of steady state force. Protein modelling studies show that these two charge alterations in alpha-TM cause a change in the surface charges of the molecule. Our results provide the first evidence that charge changes at the carboxy-terminal of alpha-TM alter the functional characteristics of the heart at both the whole organ and myofilament levels.
authors.affiliation: Cardiovascular Research Institute and Department of Medical Physiology, College of Medicine, Texas A & M University System Health Science Center, College Station, TX 77843-1114, USA. marim@tamu.edu
authors.forename: Robert D
authors.initials: RD
authors.lastname: Gaffin
authors.forename: Kuppan
authors.initials: K
authors.lastname: Gokulan
authors.forename: James C
authors.initials: JC
authors.lastname: Sacchettini
authors.forename: Timothy
authors.initials: T
authors.lastname: Hewett
authors.forename: Raisa
authors.initials: R
authors.lastname: Klevitsky
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Robbins
authors.forename: Mariappan
authors.initials: M
authors.lastname: Muthuchamy
mesh_terms: D008841:Actin Cytoskeleton / Q000502:physiology; D000818:Animals; D002118:Calcium / Q000378:metabolism; D004563:Electrochemistry; D006339:Heart Rate / Q000502:physiology; D051379:Mice; D008822:Mice, Transgenic; D016297:Mutagenesis, Site-Directed; D009200:Myocardial Contraction / Q000502:physiology*; D017434:Protein Structure, Tertiary; D014335:Tropomyosin / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D016277:Ventricular Function, Left / Q000502:physiology
chemical_list: D014335:Tropomyosin; D002118:Calcium