大鼠肾小管膜中的降钙素敏感腺苷酸环化酶。

Calcitonin-sensitive adenylate cyclase in rat renal tubular membranes.

作者信息

Loreau N, Lepreux C, Ardaillou R

出版信息

Biochem J. 1975 Sep;150(3):305-14. doi: 10.1042/bj1500305.

DOI:10.1042/bj1500305

PMID:2153

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC1165744/

Abstract

Renal tubular membranes from rat kidneys were prepared, and adenylate cyclase activity was measured under basal conditions, after stimulation by NaF or salmon calcitonin. Apparent Km value of the enzyme for hormone-linked receptor was close to 1 x 10(-8) M. 2. The system was sensitive to temperature and pH. pH was found to act both on affinity for salmon calcitonin-linked receptor and maximum stimulation, suggesting an effect of pH on hormone-receptor binding and on a subsequent step. 3. KCl was without effect areas whereas CoCl and CaCl2 above 100 muM and MnCl2 above 1 muM inhibited F- -and salmon calcitonin-sensitive adenylate cyclase activities. The Ca2+ inhibition of the response reflected a fall in maximum stimulation and not a loss of affinity of salmon calcitonin-linked receptor for the enzyme. 4. The measurement of salmon calcitonin-sensitive adenylate cyclase activity as a function of ATP concentration showed that the hormone increases the maximum velocity of the adenylate cyclase. GTP, ITP and XTP at 200 muM did not modify basal, salmon calcitonin- and parathyroid hormone-sensitive adenylate cyclase activities. 5. Basal, salmon calcitonin- and F- -sensitive adenylate cyclase activities decreased at Mg2+ concentrations below 10 mM. High concentrations of Mg2+ (100 mM) led to an inhibition of the F- -stimulated enzyme. 6. Salmon calcitonin-linked receptor had a greater affinity for adenylate cyclase than human or porcine calcitonin-linked receptors. There was no additive effect of these three calcitonin peptides whereas parathyroid hormone added to salmon calcitonin increased adenylate cyclase activity, thus showing that both hormones bound to different membrane receptors. Human calcitonin fragments had no effect on adenylate cyclase activity. 7. Salmon calcitonin-stimulated adenylate cyclase activity decreased with the preincubation time. This was due to progressive degradation of the hormone and not to the rate of binding to membrane receptors.

摘要

制备大鼠肾脏的肾小管膜，在基础条件下、经氟化钠（NaF）或鲑鱼降钙素刺激后测量腺苷酸环化酶活性。该酶与激素连接受体的表观米氏常数（Km值）接近1×10⁻⁸ M。2. 该系统对温度和pH敏感。发现pH既作用于对鲑鱼降钙素连接受体的亲和力，也作用于最大刺激，表明pH对激素-受体结合及后续步骤有影响。3. 氯化钾（KCl）无作用，而100 μM以上的氯化钴（CoCl）和氯化钙（CaCl₂）以及1 μM以上的氯化锰（MnCl₂）抑制氟离子（F⁻）和鲑鱼降钙素敏感的腺苷酸环化酶活性。钙离子对反应的抑制反映最大刺激的降低，而非鲑鱼降钙素连接受体对该酶亲和力的丧失。4. 作为三磷酸腺苷（ATP）浓度函数的鲑鱼降钙素敏感的腺苷酸环化酶活性测量表明，该激素增加了腺苷酸环化酶的最大反应速度。200 μM的鸟苷三磷酸（GTP）、肌苷三磷酸（ITP）和黄苷三磷酸（XTP）未改变基础、鲑鱼降钙素和甲状旁腺激素敏感的腺苷酸环化酶活性。5. 在镁离子（Mg²⁺）浓度低于10 mM时，基础、鲑鱼降钙素和氟离子敏感的腺苷酸环化酶活性降低。高浓度的镁离子（100 mM）导致对氟离子刺激的酶的抑制。6. 鲑鱼降钙素连接受体对腺苷酸环化酶的亲和力大于人或猪降钙素连接受体。这三种降钙素肽没有相加作用，而添加到鲑鱼降钙素中的甲状旁腺激素增加了腺苷酸环化酶活性，因此表明两种激素结合到不同的膜受体上。人降钙素片段对腺苷酸环化酶活性无影响。7. 鲑鱼降钙素刺激的腺苷酸环化酶活性随预孵育时间而降低。这是由于激素的逐渐降解，而非与膜受体的结合速率。