Arylisothiocyanate modification of sarcoplasmic Ca2+-stimulated ATPase.
The related probes phenylisothiocyanate and p-sulfophenylisothiocyanate possess comparable reactivity with nucleophiles but are dissimilar in their solubility characteristics. The reagents are utilized to topologically characterize the sites of covalent interaction with the Ca2+-stimulated ATPase of sarcoplasmic reticulum membranes. The hydrophobic probe phenylisothiocyanate binds covalently to the membrane-integrated protein. The extent of covalent interaction of this probe is reduced to a limited level of label incorporation by either preincubation with p-sulfophenylisothiocyanate or by exposing the labeled protein to alkaline reductive conditions. With respect to the chemical nature a dual interaction of phenylisothiocyanate is postulated. Phenylisothiocyanate modifies the Ca2+-ATPase hydrophobically. In addition, aqueous-exposed nucleophiles (cysteine thiols) interact with both arylisothiocyanates. Inhibition of the Ca2+-stimulated ATPase activity is effected by either probe.
D000818:Animals; D001557:Benzenesulfonates / Q000378:metabolism*; D001667:Binding, Competitive; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D002250:Carbon Radioisotopes; D007425:Intracellular Membranes / Q000201:enzymology*; D017879:Isothiocyanates; D007700:Kinetics; D009132:Muscles / Q000201:enzymology; D011485:Protein Binding; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D013861:Thiocyanates / Q000378:metabolism*
D001557:Benzenesulfonates; D002250:Carbon Radioisotopes; D017879:Isothiocyanates; D013861:Thiocyanates; C005441:phenylisothiocyanate; C015308:4-isothiocyanatobenzenesulfonate; D000252:Calcium-Transporting ATPases