Effects of calcium on phosphatidylserine- and saposin C-stimulated glucosylceramide beta-glucosidase activity.
Glucosylceramide beta-glucosidase is a membrane-bound lysosomal hydrolase that is activated by acidic lipids, the most effective of which is phosphatidylserine (PtdSer), and an activator protein, saposin C. This report documents effects of Ca2+ ions on PtdSer- and saposin C-enhanced beta-glucosidase activity. Ca2+ either increased or decreased enzyme activity, depending on (1) the concentration of phospholipid, and (2) the presence or absence of saposin C. At PtdSer concentrations between 7.6 and 76 microM, in the absence of saposin C, Ca2+ caused an increase in beta-glucosidase activity up to 3 times that measured with PtdSer alone; this was due to both an increase in Vmax, and a decrease in Km. In contrast, at PtdSer concentrations greater than 100 microM, Ca2+ inhibited beta-glucosidase activity by 50%, due to a 2-fold increase in Km. Ca2+ was inhibitory at all PtdSer concentrations tested when both PtdSer and saposin C were present in the assay. Ca2+ ions were also shown to cause changes in the aggregation states of PtdSer. These results suggest that changes in Ca2+ concentration may play a role in regulating beta-glucosidase activity in vivo, thereby modulating sphingolipid metabolism. The implications of these findings are discussed.
Prence
E M
EM
Division of Medical Genetics, E. K. Shriver Center, Waltham, MA 02254, USA.
D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D004789:Enzyme Activation; D005962:Glucosylceramidase / Q000302:isolation & purification / Q000378:metabolism*; D006023:Glycoproteins / Q000494:pharmacology*; D007425:Intracellular Membranes / Q000201:enzymology; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008247:Lysosomes / Q000201:enzymology; D010718:Phosphatidylserines / Q000494:pharmacology*; D051381:Rats; D049231:Saposins
D006023:Glycoproteins; D010718:Phosphatidylserines; D049231:Saposins; D005962:Glucosylceramidase; D002118:Calcium