title: Subcellular localization and properties of alkaline inorganic pyrophosphatase in human polymorphonuclear leucocytes.
abstract: A new rapid assay for inorganic pyrophosphatase has been developed and the procedure optimised for measurement of the enzyme in human neutrophils. Kinetic studies showed that the activity was optimal at pH 8.0 and was activated by Mg2+. No neutral or acid pyrophosphatase was detected. Neutrophils were homogenised in isotonic sucrose and, after low speed centrifugation the intracellular localization of pyrophosphatase was determined by analytical subcellular fractionation with sucrose density gradient centrifugation. Pyrophosphatase was shown to have a dual localization to mitochondria and cytosol. No activity could be attributed to either the endoplasmic reticulum or alkaline phosphatase-containing granules (phosphasomes). Inhibitor studies clearly show that the cytosolic and mitochondrial pyrophosphatases are due to distinct enzymes. Neutrophils were isolated from control subjects, patients with chronic granulocytic leukaemia and subjects in the third trimester of pregnancy. The specific activity (mU/mg protein) of pyrophosphatase, in contrast to that of alkaline phosphatase was similar in the three groups. Levamisole, a potent inhibitor of alkaline phosphatase had no effect on pyrophosphatase activity, confirming that this activity is not attributable to neutrophil alkaline phosphatase.
mesh_terms: D003600:Cytosol / Q000201:enzymology; D004072:Digitonin / Q000494:pharmacology; D005260:Female; D006801:Humans; D008928:Mitochondria / Q000201:enzymology; D009504:Neutrophils / Q000201:enzymology*; D011247:Pregnancy; D011755:Pyrophosphatases / Q000097:blood* / Q000502:physiology
chemical_list: D011755:Pyrophosphatases; D004072:Digitonin