Uncoupling of ATP splitting from Ca(2+)-transport in Ca(2+)-transporting ATPase of the sarcoplasmic reticulum as a result of modification by N-(3-pyrene)maleimide: activation of a channel with a specificity for alkaline earth metal ions.
The sarcoplasmic reticulum (SR) membranes of rabbit skeletal muscle were allowed to react with N-(3-pyrene)maleimide (PMI) at pH 7 at 30 degrees C. The Ca(2+)-transporting activity of the SR membranes was reduced to 20% when PMI was bound to the extent of 1 mol/mol of Ca(2+)-transporting ATPase. The ATPase and the E-P forming activities were not affected by the binding of PMI up to 2 mol/mol ATPase, indicating that PMI somehow uncoupled Ca(2+)-transport from ATP splitting. Permeability of the SR membranes to Ca2+ ions was increased in parallel with the loss of the Ca(2+)-transporting activity. Of several components of the SR membranes which are reactive with PMI, the ATPase protein was the only one whose modification by PMI was directly related to the loss of the Ca(2+)-transporting activity. Similar results were obtained with the light SR membrane fraction, which lacks the ryanodine receptor, a well-recognized Ca2+ channel. These results indicated that a Ca2+ channel that would have been latent or properly regulated in native ATPase somehow escaped from the normal control mechanism as a result of modification of its SH groups by PMI and went into runaway operation. The activated channel was specific for alkaline earth metal ions, so permeability to other solutes including Co2+, Ni2+, and sucrose remained unchanged after treatment with PMI.(ABSTRACT TRUNCATED AT 250 WORDS)
Suzuki
T
T
Department of Physiological Chemistry, Tokyo Metropolitan Institute of Medical Science.
Kawakita
M
M
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000187:drug effects* / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D006868:Hydrolysis; D008301:Maleimides / Q000494:pharmacology*; D008673:Metals, Alkaline Earth / Q000378:metabolism*; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000187:drug effects / Q000201:enzymology*; D013439:Sulfhydryl Reagents / Q000494:pharmacology
D015220:Calcium Channels; D008301:Maleimides; D008673:Metals, Alkaline Earth; D013439:Sulfhydryl Reagents; D000255:Adenosine Triphosphate; C003944:N-(3-pyrene)maleimide; D000252:Calcium-Transporting ATPases; D002118:Calcium