A novel method for the isolation of calsequestrin from porcine skeletal muscle sarcoplasmic reticulum.
Calsequestrin was selectively solubilized from porcine skeletal muscle sarcoplasmic reticulum by incubation with HgCl2 and deoxycholate. While deoxycholate alone caused non-specific solubilization of calsequestrin and other extrinsic proteins, HgCl2 alone failed to extract significant amounts of extrinsic proteins. The simultaneous presence of HgCl2 and deoxycholate was necessary for the selective and quantitative solubilization of calsequestrin. Following DEAE-Sephadex ion-exchange chromatography of the HgCl2/ deoxycholate-solubilized material, calsequestrin was obtained in nearly homogeneous form. Porcine calsequestrin was similar to rabbit calsequestrin with respect to its amino acid composition and apparent molecular weight as determined by SDS-polyacrylamide gel electrophoresis. The calcium-binding capacity of porcine calsequestrin was 400 nmol of Ca2+ bound/mg protein, lower than the values reported for the rabbit protein.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002155:Calsequestrin / Q000302:isolation & purification*; D002852:Chromatography, Ion Exchange; D003840:Deoxycholic Acid; D004591:Electrophoresis, Polyacrylamide Gel; D008627:Mercuric Chloride; D008628:Mercury; D008722:Methods; D009124:Muscle Proteins / Q000302:isolation & purification*; D012519:Sarcoplasmic Reticulum / Q000032:analysis*; D012995:Solubility; D013552:Swine
D000596:Amino Acids; D002155:Calsequestrin; D009124:Muscle Proteins; D003840:Deoxycholic Acid; D008627:Mercuric Chloride; D008628:Mercury