Knauf H, Sellinger M, Haag K, Wais U
Am J Physiol. 1985 Mar;248(3 Pt 2):F389-95. doi: 10.1152/ajprenal.1985.248.3.F389.
High HCO3(-)-ATPase activity is known to exist in mitochondria of renal tubular cells. In brush border membrane (BBM) preparations of proximal tubules such an anion-stimulated enzyme was also found. However, these preparations always contained mitochondrial markers. The putative localization and the role of this ATPase in BBM is still controversial. Some authors consider the HCO3(-)-ATPase in the BBM to be a mitochondrial contamination; others attribute to this ATPase a key role in H+ transport in the proximal tubule. To reinvestigate this problem, BBMs from rat kidney cortex were isolated by a simple, rapid (1.5-h) Ca2+-precipitation method, yielding a BBM fraction enriched 12.4-fold with respect to the marker enzyme leucine aminopeptidase (LAP). There was no basolateral Na+-K+-ATPase and no mitochondrial succinate dehydrogenase detectable. Cytochrome c oxidase was drastically reduced to 7 +/- 1% of that observed in the homogenate (TH). The activity of HCO3(-)-ATPase in the BBM fraction was 19 +/- 4 IU/g protein, i.e., 27% that of the homogenate. As sonication of the TH exclusively increases the activity of HCO3(-)-ATPase, its relative activity was 7.5% and thus equal to that of the mitochondrial marker. In many BBM preparations no HCO3(-)-ATPase was detectable. In those BBM preparations in which traces of HCO3(-)-ATPase were found, this activity coincided with that of cytochrome c oxidase in the respective preparation. There was a constant activity ratio of cytochrome c oxidase/HCO3(-)-ATPase in the TH, BBM, and pellet 1. The activity of HCO3(-)-ATPase in BBM did not depend on the activity of LAP.(ABSTRACT TRUNCATED AT 250 WORDS)
已知高HCO₃⁻ -ATP酶活性存在于肾小管细胞的线粒体中。在近端小管的刷状缘膜（BBM）制剂中也发现了这种阴离子刺激酶。然而，这些制剂总是含有线粒体标记物。这种ATP酶在BBM中的假定定位和作用仍存在争议。一些作者认为BBM中的HCO₃⁻ -ATP酶是线粒体污染物；另一些人则认为这种ATP酶在近端小管的H⁺转运中起关键作用。为了重新研究这个问题，通过一种简单、快速（1.5小时）的Ca²⁺沉淀法从大鼠肾皮质中分离出BBM，得到的BBM部分相对于标记酶亮氨酸氨肽酶（LAP）富集了12.4倍。未检测到基底外侧Na⁺ -K⁺ -ATP酶和线粒体琥珀酸脱氢酶。细胞色素c氧化酶急剧减少至匀浆（TH）中观察到的7±1%。BBM部分中HCO₃⁻ -ATP酶的活性为19±4 IU/g蛋白质，即匀浆的27%。由于TH的超声处理仅增加了HCO₃⁻ -ATP酶的活性，其相对活性为7.5%，因此与线粒体标记物的活性相当。在许多BBM制剂中未检测到HCO₃⁻ -ATP酶。在那些发现有微量HCO₃⁻ -ATP酶的BBM制剂中，这种活性与相应制剂中的细胞色素c氧化酶活性一致。在TH、BBM和沉淀1中细胞色素c氧化酶/HCO₃⁻ -ATP酶的活性比恒定。BBM中HCO₃⁻ -ATP酶的活性不依赖于LAP的活性。（摘要截断于250字）
