Linde A, Granström G
J Biol Buccale. 1978 Dec;6(4):293-308.
The same isoenzyme of nonspecific alkaline phosphatase (APase), assayed with p-nitrophenylphosphate (p-NPP), was shown be present in different calcifying tissues, bone, calcifying cartilage, odontoblasts and enamel organ. Indications were also found that the enzymatic degradation of inorganic pyrophosphate (PPi) in calcifying tissues is mediated by APase. By using specific APase inhibitors, it was shown that two enzymes capable of degrading ATP exist. These were characterized in dentinogenically active odontoblasts, and it was concluded that one is the classical APase, the other is a Ca2+ and Mg2+ activated ATPase, named Ca2+-ATPase. The two phosphatases were solubilized from odontoblasts and separated. The localization of APase and Ca2+-ATPase in odontoblasts was investigated by subcellular fractionation and EM histochemistry. Routine methods for fixation were found to almost completely inactivate the enzymes. By using a mild fixation technique that preserved 80% of the enzyme activity, the main localization for both APase and Ca2+-ATPase was found to be in the membranes of intercellular vesicles located in the cell body and odontoblasts process. No activity was found in the cell membranes. It is concluded that there are at least two enzymes able to degrade phosphate compounds at alkaline pH in hard tissue forming cells. One is the nonspecific alkaline phosphatase (APase; EC 3. 1. 3. 1), which is active against p-NPP, PPi, glycerophosphates and ATP among other substrates. The other is a more specific Ca2+-ATPase (EC 3. 6. 1. 3). There seems to be an intimate relation between these two enzymes in the tissue. The function of APase in biological calcification is still obscure. In contrast, the finding of an ATP dependent, intravesicularly directed, transmembranous Ca2+-transport in vesicles derived from the microsomal fraction of odontoblasts may explain the role of Ca2+-ATPase.
用对硝基苯磷酸酯（p-NPP）检测的非特异性碱性磷酸酶（APase）的同一种同工酶，在不同的钙化组织，如骨骼、钙化软骨、成牙本质细胞和釉器中均有发现。研究还发现钙化组织中无机焦磷酸（PPi）的酶促降解是由APase介导的。通过使用特异性APase抑制剂，发现存在两种能够降解ATP的酶。对具有牙本质形成活性的成牙本质细胞中的这些酶进行了表征，得出的结论是，一种是经典的APase，另一种是Ca2+和Mg2+激活的ATP酶，命名为Ca2+-ATPase。这两种磷酸酶从成牙本质细胞中溶解并分离出来。通过亚细胞分级分离和电子显微镜组织化学研究了APase和Ca2+-ATPase在成牙本质细胞中的定位。发现常规固定方法几乎能使这些酶完全失活。通过使用一种能保留80%酶活性的温和固定技术，发现APase和Ca2+-ATPase的主要定位都在位于细胞体和成牙本质细胞突起中的细胞间小泡膜上。在细胞膜上未发现活性。得出的结论是，在硬组织形成细胞中至少有两种酶能够在碱性pH下降解磷酸化合物。一种是非特异性碱性磷酸酶（APase；EC 3.1.3.1），它对p-NPP、PPi、甘油磷酸酯和ATP等其他底物具有活性。另一种是更具特异性的Ca2+-ATPase（EC 3.6.1.3）。在组织中这两种酶之间似乎存在密切关系。APase在生物钙化中的功能仍不清楚。相比之下，在源自成牙本质细胞微粒体部分的小泡中发现的依赖ATP的、泡内定向的跨膜Ca2+转运，可能解释了Ca2+-ATPase的作用。
