Effect of K+, and other ligands on the thiol reactivity and tryptic cleavage pattern of scallop sarcoplasmic reticulum.
The kinetics of reaction of the thiol groups of both membranous and non-ionic detergent-solubilized Ca-ATPase of scallop sarcoplasmic reticulum towards 5,5'-dithiobis(2-nitrobenzoate) were greatly modified in different ways by the presence of the following combinations of ligands: Ca2+, EGTA (no Ca2+), (ATPMg2- + EGTA) and (ATP + Ca2+). K+ was found to influence greatly the pattern of reactivity of the thiol groups of the scallop Ca-ATPase, modifying the kinetics of reaction differently according to the types of other ligand present. While all the thiol groups on the non-ionic detergent-solubilized Ca-ATPase were available for reaction in the absence of K+, whatever the combination of ligands, in the presence of K+, several groups became completely unreactive towards the reagent. In some cases the rate of inactivation of Ca-ATPase activity could be related to the rates of reaction of different kinetic classes of thiol group, according to the ligands present. Large differences were also seen in the tryptic cleavage pattern in the presence of the different ligands, and K+ led to major modifications in the products of digestion in the absence of nucleotide. An 80 kDa tryptic fragment was observed, as with lobster Ca-ATPase but unlike rabbit skeletal muscle Ca-ATPase. A complete amino-acid analysis of the scallop Ca-ATPase was carried out. Differences in the non-polar amino-acid content from rabbit skeletal muscle Ca-ATPase may relate to the different lipid composition of the two membranes.
Hardwicke
P M
PM
Department of Medical Biochemistry, Southern Illinois University, Carbondale 62901-4409.
Huvos
P
P
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D003902:Detergents; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology; D007700:Kinetics; D008024:Ligands; D008974:Mollusca / Q000201:enzymology*; D010766:Phosphorylation; D011188:Potassium / Q000494:pharmacology*; D011487:Protein Conformation; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D012995:Solubility; D013329:Structure-Activity Relationship; D013438:Sulfhydryl Compounds; D014357:Trypsin / Q000494:pharmacology*
D000596:Amino Acids; D003902:Detergents; D008024:Ligands; D013438:Sulfhydryl Compounds; D004228:Dithionitrobenzoic Acid; D014357:Trypsin; D000252:Calcium-Transporting ATPases; D011188:Potassium; D002118:Calcium