Sasaki T, Naka M, Nakamura F, Tanaka T
Department of Molecular and Cellular Pharmacology, Mie University School of Medicine, Japan.
J Biol Chem. 1992 Oct 25;267(30):21518-23.
The Ca(2+)-calmodulin (CaM)-dependent activation of myosin light chain kinase is inhibited by ruthenium red competitively with respect to Ca2+, with a Ki value of 8.6 microM. The binding of Ca2+ to CaM is inhibited by micromolar concentrations of ruthenium red. In the absence of Ca2+, CaM has two binding sites for ruthenium red with the dissociation constants of 0.36 and 8.7 microM, respectively. Ca2+ antagonizes the binding of ruthenium red to the low-affinity site on CaM. Binding of ruthenium red to the high-affinity site is not affected by Ca2+. The low- and high-affinity sites for ruthenium red are shown to be located in the NH2-terminal half and the COOH-terminal half of CaM, respectively. Lower concentrations of ruthenium red are needed for enzyme inactivation than for the dissociation of enzyme-CaM-Sepharose complex, suggesting these events have different Ca2+ requirements. Moreover, ruthenium red inhibits Ca(2+)-induced contraction of depolarized vascular smooth muscle in a competitive manner with respect to Ca2+. These results suggest that ruthenium red may be a new type of CaM antagonist that inhibits the binding of Ca2+ to CaM and thereby inhibits Ca(2+)-CaM-dependent enzymes and smooth muscle contraction competitively with respect to Ca2+.
钌红相对于Ca2+竞争性地抑制了肌球蛋白轻链激酶的Ca(2+)-钙调蛋白（CaM）依赖性激活，其Ki值为8.6微摩尔。微摩尔浓度的钌红抑制Ca2+与CaM的结合。在没有Ca2+的情况下，CaM有两个钌红结合位点，其解离常数分别为0.36和8.7微摩尔。Ca2+拮抗钌红与CaM上低亲和力位点的结合。钌红与高亲和力位点的结合不受Ca2+影响。钌红的低亲和力和高亲和力位点分别位于CaM的NH2末端一半和COOH末端一半。酶失活所需的钌红浓度低于酶-CaM-琼脂糖复合物解离所需的浓度，这表明这些事件对Ca2+的需求不同。此外，钌红以相对于Ca2+的竞争性方式抑制Ca(2+)诱导的去极化血管平滑肌收缩。这些结果表明，钌红可能是一种新型的CaM拮抗剂，它抑制Ca2+与CaM的结合，从而相对于Ca2+竞争性地抑制Ca(2+)-CaM依赖性酶和平滑肌收缩。
