Tryptic digestion of sarcoplasmic reticulum inhibits Ca2+ accumulation by action on a membrane component other than the Ca2+-ATPase.
We have reexamined the "uncoupling" of Ca2+ transport from ATP hydrolysis, which has been reported to be caused by trypsin cleavage of the Ca2+-ATPase of sarcoplasmic reticulum (SR) vesicles at the second (slower) of two characteristic tryptic sites (Scott, T. L., and Shamoo, A. E. (1982) J. Membr. Biol. 64, 137-144). We find that the loss of Ca2+ accumulation capacity in SR vesicles is poorly correlated with this cleavage under several conditions. The loss is accompanied by increased Ca2+ permeability but not by changes in the properties of the ATPase or ATP-Pi exchange activities of the vesicles. Proteoliposomes containing purified Ca2+-ATPase which has been cleaved in part at the two tryptic sites are as well coupled and impermeable to Ca2+ as proteoliposomes containing intact Ca2+-ATPase. We conclude that the loss of Ca2+ accumulation capacity in SR vesicles on tryptic treatment is due to cleavage of a SR membrane component other than the Ca2+-ATPase, possibly a component of the gated channels which function in Ca2+ release from SR, which leads to a Ca2+ leak. The hydrolytic and coupled transport functions of the Ca2+-ATPase itself may well be unaffected by the two tryptic cleavages.
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D008970:Molecular Weight; D011510:Proteolipids / Q000378:metabolism; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*; D014357:Trypsin / Q000378:metabolism*
D011510:Proteolipids; C020485:proteoliposomes; D014357:Trypsin; D000252:Calcium-Transporting ATPases; D002118:Calcium