A calcyclin-associated protein is a newly identified member of the Ca2+/phospholipid-binding proteins, annexin family.
A calcyclin-associated protein with an apparent molecular weight of 50,000 (CAP-50) was purified from rabbit lung. The procedure included ammonium sulfate precipitation, anion and cation ion-exchange, and calcyclin affinity chromatographies. Interestingly, partial amino acid sequences of lysyl-endpeptidase-digested fragments indicated that CAP-50 was a member of the Ca2+/phospholipid-binding proteins, the annexin family. The sequence of a proteolytic peptide with Staphylococcus aureus V8 protease on NH2-terminal region is not homologous with any other annexin family proteins. Phospholipid binding studies showed that CAP-50 bound to phosphatidylserine, phosphatidylethanolamine, phosphatidylinositol, and phosphatidic acid-containing vesicles, in a Ca(2+)-dependent manner. In the presence of Ca2+/calcyclin, CAP-50 formed a complex with calcyclin and bound to the PS-containing vesicles. The apparent Kd value of calcyclin for CAP-50 was calculated to be 1.61 x 10(-6) M. Zero-length cross-linking studies indicated that 1 mol of CAP-50 bound to an equimolar unit of calcyclin. CAP-50 inhibited the phospholipase A2 activity, dose-dependently (IC50 = 0.2 microM), however, calcyclin did not alter the inhibitory effect. With the 125I-calcyclin gel overlay method, calcyclin bound tightly to CAP-50 in a Ca(2+)-dependent manner after sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These results suggest that rabbit lung CAP-50 is a newly identified member of the annexin family. Ca2+/calcyclin apparently regulates the function of CAP-50 on cytosolic face of the plasma membrane.
Tokumitsu
H
H
Department of Pharmacology, Nagoya University School of Medicine, Japan.
Mizutani
A
A
Minami
H
H
Kobayashi
R
R
Hidaka
H
H
D000595:Amino Acid Sequence; D000818:Animals; D017302:Annexins; D001345:Autoradiography; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism*; D002413:Cations, Divalent; D002846:Chromatography, Affinity; D003432:Cross-Linking Reagents; D004591:Electrophoresis, Polyacrylamide Gel; D008168:Lung / Q000737:chemistry; D008969:Molecular Sequence Data; D005810:Multigene Family; D010743:Phospholipids / Q000378:metabolism; D011817:Rabbits; D009418:S100 Proteins; D012689:Sequence Homology, Nucleic Acid
D017302:Annexins; D002135:Calcium-Binding Proteins; D002413:Cations, Divalent; D003432:Cross-Linking Reagents; D010743:Phospholipids; D009418:S100 Proteins; C074368:calcyclin-associated protein 50; D002118:Calcium