Adsorption of sphingomyelinase of Bacillus cereus onto erythrocyte membranes.
Sphingomyelinase of Bacillus cereus proved to be specifically adsorbed onto mammalian erythrocyte membranes in the presence of either Ca2+ or Ca2+ plus Mg2+ in the order of sphingomyelin content; i.e., sheep, bovine greater than porcine greater than rat erythrocytes. No appreciable adsorption was observed in the presence of Mg2+ alone nor in the absence of divalent metal ions. The enzyme adsorption onto bovine erythrocytes was dependent upon the incubation temperature. By shifting the temperature from 37 to 0 degrees C, sphingomyelinase once adsorbed onto the surface of bovine erythrocytes was released into the supernatant. Ca2+ proved to be an essential factor for the enzyme adsorption: The addition of 1 mM Ca2+ enhanced the adsorptive process, but inhibited sphingomyelin hydrolysis and hot or hot-cold hemolysis of erythrocytes, while the addition of 1 mM Ca2+ plus 1 mM Mg2+ enhanced sphingomyelin breakdown and hemolysis as well as the enzyme adsorption. However, when the amount of sphingomyelin fell off to 0.2-0.7 nmol/ml or less by the action of sphingomyelinase, the enzyme once adsorbed was completely released from the surface of erythrocytes. The result indicates that the major binding site for sphingomyelinase is sphingomyelin. In the presence of 1 mM Mg2+ alone, the enzymatic hydrolysis of sphingomyelin and hemolysis proceeded whereas the enzyme adsorption was not encountered during 60 min incubation at 37 degrees C. The change in the molar ratio of Ca2+ to Mg2+ affected the enzyme adsorption and sphingomyelin breakdown; the higher Ca2+ enhanced the adsorption whereas the higher Mg2+ stimulated sphingomyelin hydrolysis.
D000327:Adsorption; D000818:Animals; D001409:Bacillus cereus / Q000201:enzymology*; D002118:Calcium / Q000097:blood; D002413:Cations, Divalent / Q000097:blood; D002417:Cattle; D004910:Erythrocyte Membrane / Q000378:metabolism*; D004912:Erythrocytes / Q000378:metabolism*; D007700:Kinetics; D010727:Phosphoric Diester Hydrolases / Q000097:blood*; D051381:Rats; D012756:Sheep; D013108:Sphingomyelin Phosphodiesterase / Q000097:blood*; D013109:Sphingomyelins / Q000097:blood; D013552:Swine; D013696:Temperature
D002413:Cations, Divalent; D013109:Sphingomyelins; D010727:Phosphoric Diester Hydrolases; D013108:Sphingomyelin Phosphodiesterase; D002118:Calcium