Subcellular localization and properties of adenosine triphosphatase in human polymorphonuclear leucocytes.

Magnesium-dependent adenosine triphosphatase (Mg2+-ATPase) activities wee studied in human neutrophilic polymorphonuclear leucocytes. Kinetic studies on whole leucocyte homogenates produced curvilinear kinetics suggesting the presence of at least two forms of Mg2+-ATPase. Neutrophils were homogenized in isotonic sucrose and, after low-speed centrifugation, the supernatant was subjected to analytical subcellular fractionation. Gradient fractions were assayed for Mg2+-ATPase and for principal organelle marker enzymes. Mg2+-ATPase was distributed between the plasma membrane, mitochondrial and cytosol fractions. Kinetic and inhibitor studies on Mg2+-ATPase from each localization indicated the presence of three forms of the enzyme. The plasma membrane and mitochondrial activities had a Km value of 0.2 mmol/l for ATP, whilst the Km for the cytosolic enzyme was 1.8 mmol/l. Inhibitor studies showed further differences between the three enzymes. Neutrophils were isolated from control subjects, patients with chronic granulocytic leukaemia and patients in the third trimester of pregnancy. The specific activities (mUnits/mg protein) of Mg2+-ATPase, in contrast to those of alkaline phosphatase, were similar in all three patient groups. This result, together with the fractionation experiments and inhibitor studies, strongly suggest that the ATPase is not attributable to neutrophil alkaline phosphatase.