Structural domain mapping and phosphorylation of human erythrocyte pallidin (band 4.2).
Pallidin (band 4.2) is a major protein of the human erythrocyte membrane, and plays an important but as yet undefined role in maintaining the normal shape and lifespan of the erythrocyte. The pallidin protein has been purified by a new procedure which yields a protein which is > 97% pure as judged by gel electrophoresis, while pallidin purified by our original procedure is only approx. 85% pure. The new form of the protein is unstable in physiological salt solutions. However, taking advantage of its high purity, we have used the new form of the protein to produce a structural domain map of its principal tryptic fragments. We also show that pallidin can be phosphorylated by a red-cell membrane kinase which partially co-purifies with it, and has properties similar to the catalytic subunit of cAMP-dependent kinase. Both cAMP-dependent kinase and the red-cell kinase phosphorylate the same tryptic domains on the pallidin protein. Our results show that endogenous pallidin on the red-cell membrane is a poor substrate for the kinase, possibly because it is fully phosphorylated, or inaccessible to the kinase.
Dotimas
E
E
Dept. of Biomedical Research, St. Elizabeth's Hospital of Boston, MA 02135.
Speicher
D W
DW
GuptaRoy
B
B
Cohen
C M
CM
D000199:Actins / Q000302:isolation & purification; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D002352:Carrier Proteins; D004912:Erythrocytes / Q000737:chemistry*; D006801:Humans; D037102:Lectins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D008565:Membrane Proteins / Q000737:chemistry; D008969:Molecular Sequence Data; D015394:Molecular Structure; D010449:Peptide Mapping; D010766:Phosphorylation; D011494:Protein Kinases / Q000032:analysis; D013049:Spectrin / Q000302:isolation & purification; D013379:Substrate Specificity; D014357:Trypsin
D000199:Actins; D000596:Amino Acids; C400284:BLOC1S6 protein, human; D002352:Carrier Proteins; D037102:Lectins; D008565:Membrane Proteins; D013049:Spectrin; D011494:Protein Kinases; D014357:Trypsin