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- W2087989672 abstract "Recombinant human cardiac actin mutants E99K, A331P, and WT were expressed in baculovirus/insect cells and used to reconstitute the thin-filament of bovine cardiac fibers, together with tropomyosin (Tm) and troponin (Tn). Effects of [Ca2+], [ATP], [phosphate] and [ADP] on tension and its transients were studied at 25°C. After actin filament reconstitution, tension of WT actin reconstituted fibers reached 0.75±0.06 T0 (T0: tension of native fibers), which is comparable to rabbit skeletal actin reconstituted fibers (0.73±0.07 T0). However, tension was less in E99K (0.60±0.08 T0) and A331P (0.55±0.03 T0). After Tm and Tn reconstitution, tension for E99K was 0.76±0.01 T0, and A331P 0.65±0.02 T0, which were less than WT (0.85±0.06 T0). E99K increased pCa50 (5.78±0.03) but decreased cooperativity (1.78±0.18) compared with WT (5.70±0.03, 2.6±0.3, respectively). A331P decreased pCa50 (5.57±0.03). E99K increased TLC (tension at pCa 8.0: 0.22±0.03 THC) compared to WT (0.10±0.02 THC), but A331P maintained similar TLC (0.08±0.01 THC) to WT. Five equilibrium constants of the cross-bridge cycle were deduced using sinusoidal analysis. E99K caused a decrease in both K0 (ADP dissociation: 16.6±4.6 mM−1) and K1 (ATP association: 1.51±0.08 mM−1) compared to WT (35.04±6.3 mM−1 and 2.1±0.3 mM−1, respectively). A331P and WT showed similar K0 (31±5.4 mM−1 and 35.04±6.3 mM−1), K1 (1.6±0.2 mM−1 and 2.1±0.3 mM−1), K2 (cross-bridge detachment: 0.69±0.1 and 0.78±0.13), K4 (force generation: 0.27±0.03 and 0.34±0.03), and K5 (Pi association: 0.13±0.2 mM−1 and 0.12±0.01 mM−1). The cross-bridge distribution was similar among WT, E99K and A331P, indicating that force/cross-bridge is decreased in K99K and A331P. In conclusion, E99K actin causes HCM through impaired relaxation and elevated pCa50 (diastolic problem), and lowered force/cross-bridge (systolic problem), whereas A331P actin causes HCM through decreased pCa50 and force/cross-bridge (systolic problem)." @default.
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- W2087989672 title "The Effect of α-Actin Mutant E361G on Force Generation and Cross-Bridge Kinetics in Thin-Filament Reconstituted Bovine Cardiac Muscle Fibers" @default.
- W2087989672 doi "https://doi.org/10.1016/j.bpj.2013.11.4248" @default.
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