Purification of phospholamban, a 22,000 dalton protein from cardiac sarcoplasmic reticulum that is specifically phosphorylated by cyclic AMP-dependent protein kinase

Technical Report · Mon Jan 01 04:00:00 EST 1979

DOI:https://doi.org/10.2172/5008458· OSTI ID:5008458

Bidlack, J. M.; Shamoo, A. E.

Very low concentrations deoxycholate (DOC) were used to isolate two proteins from canine cardiac sarcoplasmic reticulum. These two proteins are phospholamban, a 22,000 dalton protein, and the Ca/sup 2 +/ + Mg/sup 2 +/-ATPase, the major protein of the sarcoplasmic reticulum, responsible for the active transport of calcium. The 22,000 dalton protein is first solubilized in a very low concentration of DOC and then subjected to column chromatography. After molecular weight sieving on a Sephadex G-75 column, the 22,000 dalton protein appears as a purified protein on sodium dodecyl sulfate (SDS)-polyacrylamide gels. The purified protein is specifically phosphorylated by cyclic AMP-dependent protein kinase. Phospholipids are still bound to the isolated protein. The Ca/sup 2 +/ + Mg/sup 2 +/-ATPase is purified by first solubilizing all the extrinsic proteins with a low concentration of DOC. An increasing amount of DOC is then added to yield the purified Ca/sup 2 +/ + Mg/sup 2 +/-ATPase. This protein is at least 95% pure. Adding additional DOC to the purified enzyme enhances the enzyme's ability to hydrolyze ATP. (ERB)

• Research Organization: Rochester Univ., NY (USA). Dept. of Radiation Biology and Biophysics
• DOE Contract Number: AC02-76EV03490
• OSTI ID: 5008458
• Report Number(s): UR-3490-1805
• Country of Publication: United States
• Language: English

Related Subjects

550200* -- Biochemistry
59 BASIC BIOLOGICAL SCIENCES
ADDITIVES
ANIMALS
ATP-ASE
BODY
CARDIOVASCULAR SYSTEM
CHEMICAL REACTIONS
CHROMATOGRAPHY
DATA
DETERGENTS
DOGS
ELECTROPHORESIS
EMULSIFIERS
ENZYME ACTIVITY
ENZYMES
ESTERASES
ESTERS
EXPERIMENTAL DATA
HEART
HYDROLASES
IN VITRO
INFORMATION
LIPIDS
LIQUID COLUMN CHROMATOGRAPHY
MAMMALS
MUSCLES
NUMERICAL DATA
ORGANIC COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
ORGANS
PHOSPHATASES
PHOSPHOLIPIDS
PHOSPHORYLATION
PHOSPHOTRANSFERASES
PROTEINS
PURIFICATION
SEPARATION PROCESSES
SURFACTANTS
TRANSFERASES
VERTEBRATES
WETTING AGENTS