The thin filament extraction and reconstitution protocol was used to investigate the functional roles of tropomyosin (Tm) isoforms and phosphorylation in bovine myocardium. The thin filament was extracted by gelsolin, reconstituted with G-actin, and further reconstituted with cardiac troponin together with one of three Tm varieties: phosphorylated alphaTm (alphaTm.P), dephosphorylated alphaTm (alphaTm.
View Article and Find Full Text PDFThe influence of Ca2+ dissociation rate from TnC and decreased cross-bridge detachment rate on the time course of relaxation induced by flash photolysis of diazo-2 in rabbit skinned psoas fibers was investigated at 15 degrees C. A TnC mutant (M82Q TnC) that exhibited increased Ca2+ sensitivity caused by a decreased Ca2+ dissociation rate in solution also increased the Ca2+ sensitivity of force and decreased the rate of relaxation in fibers approximately 2-fold. In contrast, a TnC mutant (NHdel TnC) with decreased Ca2+ sensitivity caused by an increased Ca2+ dissociation rate in solution decreased Ca2+ sensitivity of force but did not accelerate relaxation.
View Article and Find Full Text PDFPolycystin-L (PCL) is an isoform of polycystin-2, the product of the second gene associated with autosomal dominant polycystic kidney disease, and functions as a Ca(2+)-regulated nonselective cation channel. We recently demonstrated that polycystin-2 interacts with troponin I, an important regulatory component of the actin microfilament complex in striated muscle cells and an angiogenesis inhibitor. In this study, using the two-microelectrode voltage-clamp technique and Xenopus oocyte expression system, we showed that the calcium-induced PCL channel activation is substantially inhibited by the skeletal and cardiac troponin I (60% and 31% reduction, respectively).
View Article and Find Full Text PDFTroponin C (TnC) is the Ca(2+)-binding subunit of the troponin complex of vertebrate skeletal muscle. It consists of two structurally homologous domains, N and C, connected by an exposed alpha-helix. The C-domain has two high-affinity sites for Ca(2+) that also bind Mg(2+), whereas the N-domain has two low-affinity sites for Ca(2+).
View Article and Find Full Text PDFThe influence of Ca(2+)-activated force, the rate of dissociation of Ca(2+) from troponin C (TnC) and decreased crossbridge detachment rate on the time course of relaxation induced by flash photolysis of diazo-2 in rabbit skinned psoas fibres was investigated at 15 degrees C. The rate of relaxation increased as the diazo-2 chelating capacity (i.e.
View Article and Find Full Text PDFThe goal of this study was to examine the mechanism of magnesium binding to the regulatory domain of skeletal troponin C (TnC). The fluorescence of Trp(29), immediately preceding the first calcium-binding loop in TnC(F29W), was unchanged by addition of magnesium, but increased upon calcium binding with an affinity of 3.3 microm.
View Article and Find Full Text PDFIn contrast to skeletal muscle, the efficiency of the contractile apparatus of cardiac tissue has long been known to be severely compromised by acid pH as in the ischemia of myocardial infarction and other cardiac myopathies. Recent reports (Westfall, M. V.
View Article and Find Full Text PDFCa2+ and human cardiac troponin I (cTnI) peptide binding to human cardiac troponin C (cTnC) have been investigated with the use of 2D [1H,15N] HSQC NMR spectroscopy. The spectral intensity, chemical shift, and line-shape changes were analyzed to obtain the dissociation ( K(D)) and off-rate ( k(off)) constants at 30 degrees C. The results show that sites III and IV exhibit 100-fold higher Ca2+ affinity than site II ( K(D(III,IV)) approximately 0.
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