Perrild H, Frost J, Robertson W R
Histochem J. 1984 Jan;16(1):71-83. doi: 10.1007/BF01003437.
A quantitative cytochemical assay for NAD+ kinase-like activity in the guinea-pig thyroid gland is described. The NADP+ produced by the activity of the kinase was used to drive the NADP+-dependent enzyme glucose-6-phosphate dehydrogenase which is endogenous to the tissue. The activity of glucose-6-phosphate dehydrogenase is greatly in excess of that of the kinase and was unaffected by the constituents of the kinase incubation medium (ATP, Mg2+ and NAD+) either alone or in combination. Kinase activity was dependent both on ATP and Mg2+, with maximal activity seen when the Mg-ATP ratio was between 1:1 and 4:1. Free ATP inhibited the activity of the enzyme. Enzyme activity was exhibited over a broad pH range (7-9) with a peak at pH 8.2. The sulphydryl-blocking agents, p-chloromercuribenzoate, iodoacetate and iodoacetamide (at 1 mM), completely abolished kinase activity but were without effect on glucose-6-phosphate dehydrogenase activity. N-ethylmaleimide and citrate (both at 1 mM) had no effect on either kinase or glucose-6-phosphate dehydrogenase activities.
本文描述了一种用于检测豚鼠甲状腺中NAD⁺激酶样活性的定量细胞化学分析方法。激酶活性产生的NADP⁺用于驱动组织内源性的依赖NADP⁺的酶葡萄糖-6-磷酸脱氢酶。葡萄糖-6-磷酸脱氢酶的活性大大超过激酶的活性，并且不受激酶孵育培养基成分（ATP、Mg²⁺和NAD⁺）单独或组合使用的影响。激酶活性既依赖于ATP也依赖于Mg²⁺，当Mg-ATP比例在1:1至4:1之间时观察到最大活性。游离ATP抑制该酶的活性。酶活性在较宽的pH范围（7 - 9）内表现出来，在pH 8.2时达到峰值。巯基阻断剂对氯汞苯甲酸、碘乙酸和碘乙酰胺（1 mM）完全消除了激酶活性，但对葡萄糖-6-磷酸脱氢酶活性没有影响。N-乙基马来酰亚胺和柠檬酸盐（均为1 mM）对激酶或葡萄糖-6-磷酸脱氢酶活性均无影响。
