Similarity of the Active Site of Phosphorylation of the Adenosine Triphosphatase for Transport of Sodium and Potassium Ions in Kidney to That for Transport of Calcium Ions in the Sarcoplasmic Reticulum of Muscle

作者：Françoise Bastide, Gerhard Meissner, Sidney Fleischer, Robert L. Post · 发表于：Journal of Biological Chemistry · 年份：1973 · DOI：10.1016/s0021-9258(19)43145-1 · 被引用次数：209 · 研究领域：Ion channel regulation and function、Mass Spectrometry Techniques and Applications、Muscle metabolism and nutrition

Abstract The sites of phosphorylation of two ion pump proteins were compared. In this usage, active means the site of covalent attachment of the phosphate group to the protein. The site of the sodium and potassium ion transport adenosine triphosphatase of guinea pig kidney was compared with that of the calcium ion transport adenosine triphosphatase of the sarcoplasmic reticulum of rabbit skeletal muscle. Each enzyme was phosphorylated by radioactive adenosine triphosphate under its unique experimental conditions. Subsequently, the two phosphoenzymes were treated identically. Each phosphoenzyme was denatured with acid and digested to a limit radioactive phosphopeptide with pronase. The two limit phosphopeptides were indistinguishable with respect to (a) pH hydrolysis profile, (b) electrophoretic mobility at pH levels 2, 3, 4, 6, and 8, (c) oxidation by periodate, (d) lack of a positive charge below neutral pH as a result of acetylation of the phosphoenzyme before digestion, and (e) insensitivity to carboxypeptidase B. Furthermore, this limit phosphopeptide was released also from both phosphoenzymes by sequential digestion with papain first and then trypsin. Interpretation of these results suggests an site tripeptide for each enzyme, seryl or threonyl, phosphoaspartyl lysine. Both phosphoenzymes also contained a cysteine near the site on the NH2-terminal side. Digestion of each radioactive phosphoenzyme with pepsin showed clear differences between the enzymes with respect to ra...