The apparent second-order rate constants (k+T) of ATP-induced cross-bridge detachment from rigor in the absence of Ca2+ were determined with laser flash photolysis of caged ATP (cATP) in alpha-toxin-permeabilized tonic, rabbit femoral artery and phasic, rabbit bladder smooth muscles. The potential effect of cATP binding to actomyosin (AM) on cross-bridge kinetics was examined by varying the initial concentration of cATP 2-fold. For a given [ATP] released from either 10 or 5 mM cATP, the kinetics of relaxation were not significantly different; the estimated dissociation constant for cATP binding to smooth muscle AM was 1-3 mM. 2. k+T was significantly higher ((9.5 +/- 1.3) x 10(4) M-1 s-1) in the phasic than in the tonic ((3.0 +/- 1.0) x 10(4) M-1 s-1) smooth muscle. 3. We conclude that the combination of the significantly lower (approximately 3 times) apparent second-order rate constant of MgATP association with the approximately 5 times higher affinity of cross-bridges for MgADP in tonic, than in phasic, smooth muscle is a major determinant of the slower kinetics of relaxation and, probably, shortening velocity of tonic smooth muscle.
摘要
在无Ca2+条件下，利用α - 毒素通透的强直性兔股动脉平滑肌和相位性兔膀胱平滑肌中笼化ATP（cATP）的激光闪光光解，测定了ATP诱导的横桥从强直状态解离的表观二级速率常数（k+T）。通过将cATP的初始浓度变化2倍，研究了cATP与肌动球蛋白（AM）结合对横桥动力学的潜在影响。对于从10 mM或5 mM cATP释放的给定[ATP]，松弛动力学无显著差异；cATP与平滑肌AM结合的估计解离常数为1 - 3 mM。2. 相位性平滑肌中的k+T（(9.5 ± 1.3) x 10(4) M-1 s-1）显著高于强直性平滑肌中的（(3.0 ± 1.0) x 10(4) M-1 s-1）。3. 我们得出结论，在强直性平滑肌中，MgATP结合的表观二级速率常数显著更低（约3倍），同时横桥对MgADP的亲和力比相位性平滑肌中高约5倍，这两者的结合是强直性平滑肌松弛动力学较慢以及可能缩短速度较慢的主要决定因素。
