Dicyclohexylcarbodiimide interaction with sarcoplasmic reticulum. Inhibition of Ca2+ efflux.
Dicyclohexylcarbodiimide (DCCD), a hydrophobic carboxyl reagent, inhibited Ca2+ release from Ca2+-loaded sarcoplasmic reticulum vesicles, induced by elevated pH, tetraphenylboron, ATP + Pi, or membrane modification with acetic anhydride. Under the conditions used, the same concentrations of DCCD were required for inhibition of Ca2+ release, Ca2+-ATPase activity, and Ca2+ uptake. On the other hand, free Ca2+ or alkaline pH prevented the inhibition by DCCD of Ca2+-ATPase and coupled Ca2+ transport but not that of Ca2+ release. Moreover, several hydrophilic carboxyl reagents inhibited Ca2+-ATPase but not Ca2+ release. We suggest that a carboxyl residue(s), located in a hydrophobic region of a protein(s), is involved in the control of Ca2+ release, where DCCD interaction with this group blocks Ca2+ release. This group is distinct from the one involved in the inhibition of Ca2+-ATPase. DCCD also inhibited [3H]ryanodine binding to junctional sarcoplasmic reticulum membranes. The presence of Ca2+ or an alkaline pH only slightly affects the degree of inhibition of ryanodine binding by DCCD. Incubation of the membranes with [14C]DCCD resulted in labeling of 350-, 170-, 140-, 53-, and 30-kDa proteins in addition to the Ca2+-ATPase. The involvement of one or all of the DCCD-labeled proteins in Ca2+ release and ryanodine binding is discussed.
Argaman
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Department of Biology, Ben Gurion University of the Negev, Beer Sheva, Israel.
Shoshan-Barmatz
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D000089:Acetic Anhydrides / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D002234:Carbodiimides / Q000494:pharmacology*; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology*; D006863:Hydrogen-Ion Concentration; D007555:Isoxazoles / Q000494:pharmacology; D011804:Quinolines / Q000494:pharmacology; D011817:Rabbits; D012433:Ryanodine / Q000378:metabolism; D012519:Sarcoplasmic Reticulum / Q000187:drug effects* / Q000378:metabolism; D013775:Tetraphenylborate / Q000494:pharmacology
D000089:Acetic Anhydrides; D002234:Carbodiimides; D007555:Isoxazoles; D011804:Quinolines; D012433:Ryanodine; C031800:acetic anhydride; C009686:N-ethyl-5-phenylisoxazolium-3'-sulfonate; D013775:Tetraphenylborate; D004024:Dicyclohexylcarbodiimide; C000470:EEDQ; D000252:Calcium-Transporting ATPases; D002118:Calcium