Mode of activation of bovine brain inositol 1,4,5-trisphosphate 3-kinase by calmodulin and calcium.
Mode of activation of bovine brain inositol 1,4,5-trisphosphate 3-kinase by calmodulin and calcium.
复制标题
钙调蛋白和钙激活牛脑肌醇 1,4,5-三磷酸 3-激酶的模式。
作者：
G. Li;M. Comte;C. Wollheim;J. Cox
根据通常用于其他钙调素刺激系统的顺序平衡模型，定量和解释了Ca~（2+）和钙调素（CaM）对纯化的牛脑Ins（1，4，5）P3激酶激活的影响。可以得出两个主要结论。(i)CaM.Ca3和CaM.Ca4一起是体外生物活性物质，与大多数其他钙调蛋白靶点一样。(ii)这些物质以非合作方式与酶结合，亲和力常数为8.23 x 10（9）M-1，即比大多数钙调蛋白激活的靶酶高约10倍。蜂毒肽和三氟拉嗪对钙调蛋白激活Ins（1，4，5）P3激酶的剂量-反应曲线没有显著影响，而在非常相似的测定条件下，当测定培养基中存在1 μ M蜂毒肽或20 μ M-三氟拉嗪时，牛脑环AMP磷酸二酯酶的半数最大激活需要高出30 - 50倍的CaM浓度。类似地，1 μ M的抗钙调蛋白肽β-纤溶酶和短杆菌肽S以及20 μ M的N-（6-氨基己基）-5-氯-1-萘磺酰胺（W7）不抑制活化过程。这些数据表明，结合和激活的Ins（1，4，5）P3激酶需要钙调素的表面位点，这是不同于那些参与结合的大多数其他靶酶或模型肽。
The effect of Ca2+ and calmodulin (CaM) on the activation of purified bovine brain Ins(1,4,5)P3 kinase was quantified and interpreted according to the model of sequential equilibria generally used for other calmodulin-stimulated systems. Two main conclusions can be drawn. (i) CaM.Ca3 and CaM.Ca4 together are the biologically active species in vitro, as is the case for the great majority of other calmodulin targets. (ii) These species bind in a non-co-operative way to the enzyme with an affinity constant of 8.23 x 10(9) M-1, i.e. approx 10-fold higher than for most calmodulin-activated target enzymes. The dose-response curve of the activation of Ins(1,4,5)P3 kinase by calmodulin is not significantly impaired by melittin and trifluoperazine, whereas under very similar assay conditions the half-maximal activation of bovine brain cyclic AMP phosphodiesterase requires over 30-50-fold higher concentrations of CaM when 1 microM melittin or 20 microM-trifluoperazine is present in the assay medium. Similarly, 1 microM of the anti-calmodulin peptides seminalplasmin and gramicidin S, as well as 20 microM of N-(6-aminohexyl)-5-chloro-1-naphthalene-sulphonamide (W7), do not inhibit the activation process. These data suggest that binding and activation of Ins(1,4,5)P3 kinase require surface sites of calmodulin which are different from those involved in the binding of most other target enzymes or of model peptides.