A Comparative Study of (Ca~(2+)-Mg~(2+))-ATPase on the Lysosomal Membrane and ecto-ATPase on the Plasma Membrane from Rat Liver

摘要

We showed that rat liver lysosomes possess at least two types of ATPase besides H+-translocating ATPase (H+-ATPase); namely, N-ethylmaleimide (NEM)-sensitive and bafilomyciii Arinsensitive Mg2+-ATPase [ATPase I] and NEM- and bafllomycin A,-insensitive (Ca2+-Mg2+)-ATPase [ATPase II] [Hayashi H., et ah Chew. P/iann. Bull., 37, 2783—2786 (1992)]. Subcellular fractionation showed the presence of similar activity of (Ca2 +-Mg2 +)-ATPase not only in the Golgi but also in the plasma membrane fractions, of which plasma membrane had the highest activity, most probably due to the ecto-ATPase. In this study, we partially purified the (Ca2+-Mg2+)-ATPases from both lysosomal and plasma membranes and compared their properties. Both enzymes had quite similar characteristics including: (1) broad pH-activity profiles with two pH optima at 4.5 and 7.0, (2) Km values for ATP being 21—27 fiM (at pH 4.5) and 18—14 ftM (at pH 7.0) (in the presence of CaCl2), (3) hydrolysis of both nucleoside triphosphates and diphosphatc (ADP), (4) inhibition only by vanadate and 4,4'-diisothiocyanatostilbene 2,2'-disulfonic acid (DIDS) at pH 4.0 (but not at pH 7.0), (5) acidic pi values that were shifted by neuraminidase digestion, and (6) a positive reaction against an antibody to the /V-terminal peptide of ecto-ATPase.