A Ca2+-dependent tryptic cleavage site and a protein kinase A phosphorylation site are present in the Ca2+ regulatory domain of scallop muscle Na+-Ca2+ exchanger.
Digestion of scallop muscle membrane fractions with trypsin led to release of soluble polypeptides derived from the large cytoplasmic domain of a Na(+)-Ca(2+) exchanger. In the presence of 1 mm Ca(2+), the major product was a peptide of approximately 37 kDa, with an N terminus corresponding to residue 401 of the NCX1 exchanger. In the presence of 10 mm EGTA, approximately 16- and approximately 19-kDa peptides were the major products. Polyclonal rabbit IgG raised against the 37-kDa peptide also bound to the 16- and 19-kDa soluble tryptic peptides and to a 105-110-kDa polypeptide in the undigested membrane preparation. The 16-kDa fragment corresponded to the N-terminal part of the 37-kDa peptide. The conformation of the precursor polypeptide chain in the region of the C terminus of the 16-kDa tryptic peptide was thus altered by the binding of Ca(2+). Phosphorylation of the parent membranes with the catalytic subunit of protein kinase A and [gamma-(32)P]ATP led to incorporation of (32)P into the 16- and 37-kDa soluble fragments. A site may exist within the Ca(2+) regulatory domain of a scallop muscle Na(+)-Ca(2+) exchanger that mediates direct modulation of secondary Ca(2+) regulation by cAMP.
Chen
M
M
Department of Biochemistry and Molecular Biology, Southern Illinois University, Carbondale, Illinois 62901-4413, USA.
Zhang
Z
Z
Tawiah-Boateng
M A
MA
Hardwicke
P M
PM
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D003902:Detergents; D006868:Hydrolysis; D008969:Molecular Sequence Data; D008974:Mollusca; D009132:Muscles / Q000378:metabolism*; D010449:Peptide Mapping; D010766:Phosphorylation; D011817:Rabbits; D017386:Sequence Homology, Amino Acid; D019831:Sodium-Calcium Exchanger / Q000737:chemistry / Q000378:metabolism*; D012995:Solubility; D014357:Trypsin / Q000378:metabolism*
D003902:Detergents; D019831:Sodium-Calcium Exchanger; D017868:Cyclic AMP-Dependent Protein Kinases; D014357:Trypsin; D002118:Calcium