Demonstration of two distinct calcium pumps in human platelet membrane vesicles.
Membrane vesicles from human platelets were prepared by various disruption and isolation techniques reported in the literature to yield fractions of predominantly surface or intracellular membrane origin. ATP + Mg2+-dependent Ca2+ accumulation and the formation of acylphosphate intermediates of the calcium pump(s) were followed in parallel experiments, and the consequences of a limited proteolysis of the membranes examined. In all types of preparations active Ca2+ uptake had both oxalate-sensitive and insensitive fractions and calmodulin had no effect on the rate of Ca2+ uptake. Limited proteolysis by trypsin eliminated oxalate-sensitive Ca2+ uptake while it had no effect on the oxalate-insensitive fraction. The Ca2+-induced EP complex had an apparent molecular mass of 100-110 kDa in all of the preparations, the EP showing a broad or even duplicated line in most autoradiographies. Mild trypsin digestion resulted in the formation of 80-, 55-, and 35-kDa phosphorylated fragments. The 80-kDa fragment corresponded to the limit polypeptide found in the proteolyzed erythrocyte membrane Ca2+ pump, its phosphorylation was stimulated by lanthanum, and it appeared in a different time course than the smaller fragments. The molecular mass and the formation pattern of the latter species corresponded to the tryptic fragments in the sarcoplasmic reticulum Ca2+ pump. Based on these results we suggest that platelet membrane preparations contain two types of Ca2+ pump proteins, one similar to the sarcoplasmic reticulum-type and the other to the erythrocyte-type enzyme.
D000251:Adenosine Triphosphatases / Q000097:blood; D000255:Adenosine Triphosphate / Q000494:pharmacology; D001792:Blood Platelets / Q000378:metabolism* / Q000648:ultrastructure; D002118:Calcium / Q000097:blood* / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D006801:Humans; D007473:Ion Channels / Q000187:drug effects / Q000378:metabolism*; D007811:Lanthanum / Q000494:pharmacology; D008274:Magnesium / Q000494:pharmacology; D008854:Microscopy, Electron; D010070:Oxalates / Q000494:pharmacology; D019815:Oxalic Acid; D010710:Phosphates / Q000097:blood; D014357:Trypsin / Q000494:pharmacology
D007473:Ion Channels; D010070:Oxalates; D010710:Phosphates; D007811:Lanthanum; D000255:Adenosine Triphosphate; D019815:Oxalic Acid; D014357:Trypsin; D000251:Adenosine Triphosphatases; D008274:Magnesium; D002118:Calcium