title: Conformational responses of the tryptic cleavage products of the Ca2+-ATPase of sarcoplasmic reticulum.
abstract: Trypsin cleaves the Ca2+-ATPase of sarcoplasmic reticulum into two major fragments (A and B), followed by subsequent cleavage into smaller peptides. Although the ATP-dependent Ca2+ transport is still observed after cleavage of the ATPase into the A and B fragments, the Ca2+ transport energized by acetyl phosphate is strongly inhibited. Covalent labeling of the Ca2+-ATPase by fluorescein 5'-isothiocyanate inhibited both the ATP and acetyl phosphate-dependent Ca2+ transport. Vanadate protected the A and B fragments from further hydrolysis and preserved the ability of the cleaved Ca2+-ATPase to form crystals and to show the characteristic conformational changes in response to Ca2+ and EGTA that are observed with the intact enzyme. The protective effect of vanadate may be useful for the isolation of the A and B fragments in functional form.
authors.forename: L
authors.initials: L
authors.lastname: Dux
authors.forename: S
authors.initials: S
authors.lastname: Papp
authors.forename: A
authors.initials: A
authors.lastname: Martonosi
mesh_terms: D000818:Animals; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000032:analysis*; D004533:Egtazic Acid / Q000494:pharmacology; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins; D066298:In Vitro Techniques; D011487:Protein Conformation; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D013861:Thiocyanates; D014357:Trypsin / Q000494:pharmacology; D014638:Vanadates; D014639:Vanadium / Q000494:pharmacology
chemical_list: D005452:Fluoresceins; D013861:Thiocyanates; D014639:Vanadium; D014638:Vanadates; D004533:Egtazic Acid; D014357:Trypsin; D000252:Calcium-Transporting ATPases; D016650:Fluorescein-5-isothiocyanate; D002118:Calcium