title: Myosin cross-bridges do not form precise rigor bonds in hypertrophic heart muscle carrying troponin T mutations.
abstract: Distribution of orientations of myosin was examined in ex-vivo myofibrils from hearts of transgenic (Tg) mice expressing Familial Hypertrophic Cardiomyopathy (FHC) troponin T (TnT) mutations I79N, F110I and R278C. Humans are heterozygous for sarcomeric FHC mutations and so hypertrophic myocardium contains a mixture of the wild-type (WT) and mutated (MUT) TnT. If mutations are expressed at a low level there may not be a significant change in the global properties of heart muscle. In contrast, measurements from a few molecules avoid averaging inherent in the global measurements. It is thus important to examine the properties of only a few molecules of muscle. To this end, the lever arm of one out of every 60,000 myosin molecules was labeled with a fluorescent dye and a small volume within the A-band (~1 fL) was observed by confocal microscopy. This volume contained on average 5 fluorescent myosin molecules. The lever arm assumes different orientations reflecting different stages of acto-myosin enzymatic cycle. We measured the distribution of these orientations by recording polarization of fluorescent light emitted by myosin-bound fluorophore during rigor and contraction. The distribution of orientations of rigor WT and MUT myofibrils was significantly different. There was a large difference in the width and of skewness and kurtosis of rigor distributions. These findings suggest that the hypertrophic phenotype associated with the TnT mutations can be characterized by a significant increase in disorder of rigor cross-bridges.
authors.affiliation: Dept of Molecular Biology & Immunology and Center for Commercialization of Fluorescence Technologies, University of North Texas, Health Science Center, 3500 Camp Bowie Blvd, Fort Worth, TX 76107, United States.
authors.forename: K
authors.initials: K
authors.lastname: Midde
authors.forename: V
authors.initials: V
authors.lastname: Dumka
authors.forename: J R
authors.initials: JR
authors.lastname: Pinto
authors.forename: P
authors.initials: P
authors.lastname: Muthu
authors.forename: P
authors.initials: P
authors.lastname: Marandos
authors.forename: I
authors.initials: I
authors.lastname: Gryczynski
authors.forename: Z
authors.initials: Z
authors.lastname: Gryczynski
authors.forename: J D
authors.initials: JD
authors.lastname: Potter
authors.forename: J
authors.initials: J
authors.lastname: Borejdo
mesh_terms: D000818:Animals; D024741:Cardiomyopathy, Hypertrophic, Familial / Q000235:genetics* / Q000473:pathology / Q000503:physiopathology; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D009129:Muscle Tonus / Q000235:genetics; D009154:Mutation / Q000235:genetics*; D009206:Myocardium / Q000378:metabolism* / Q000473:pathology*; D009210:Myofibrils / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D009218:Myosins / Q000378:metabolism*; D020107:Troponin T / Q000235:genetics*
chemical_list: D020107:Troponin T; D009218:Myosins