Bayley P, Ahlström P, Martin S R, Forsen S
Biochem Biophys Res Commun. 1984 Apr 16;120(1):185-91. doi: 10.1016/0006-291x(84)91431-1.
The rate of calcium dissociation from bovine testis calmodulin was measured by fluorescence stopped-flow using the calcium indicator Quin 2 or the fluorescence probe 8-anilinonaphthalene sulphonate. Two processes are resolved with Quin 2 corresponding to dissociation from the high affinity sites (kdiss 2 to 9 s-1 for T = 11 to 28 degrees C) and from the low affinity sites (kdiss 293 to 550 s-1 for T = 11 to 19 degrees C). These rates and the activation parameters as determined for the slow process delta H not equal to = 59 +/- 10 kJ X mol-1 and delta S not equal to = 30 +/- 30 JK-1 X mol-1 are in good agreement with values determined from the 43Ca NMR exchange rates. These experiments provide confirmation that the calcium induced conformational change cannot be resolved kinetically from the calcium binding or dissociation, and by inference this conformational change is not a rate-limiting process in the function of calmodulin.
使用钙指示剂喹啉-2（Quin 2）或荧光探针8-苯胺基萘磺酸盐，通过荧光停流法测定了牛睾丸钙调蛋白中钙的解离速率。用喹啉-2分辨出两个过程，分别对应于从高亲和力位点的解离（在11至28℃时，解离常数kdiss为2至9 s-1）和从低亲和力位点的解离（在11至19℃时，kdiss为293至550 s-1）。这些速率以及为慢过程确定的活化参数，即焓变ΔH≠ = 59±10 kJ·mol-1和熵变ΔS≠ = 30±30 J·K-1·mol-1，与从43Ca NMR交换速率确定的值高度一致。这些实验证实，钙诱导的构象变化在动力学上无法与钙的结合或解离区分开来，由此推断，这种构象变化在钙调蛋白的功能中不是一个限速过程。
