Identification of calmodulin-sensitive Ca(2+)-transporting ATPase in the plasma membrane of bovine corneal epithelial cell.
ATP-dependent Ca2+ uptake was characterized in a plasma membrane enriched fraction obtained from the bovine corneal epithelium. This uptake essentially represented intravesicular accumulation because 72% of the Ca2+ content was releasable following exposure to 10(-6) M A23187. The substrate and Ca2+ requirements for maximal transport activity were similar to those described in the red blood cell because: (1) exogenous calmodulin (3 microM) significantly decreased the apparent Km for Ca2+ to 0.31 microM and increased the rate of Ca2+ uptake; (2) a hydroxylamine labile Ca(2+)-dependent phosphoenzyme intermediate was identified with an apparent molecular size of 140 kDa; (3) Ca(2+)-dependent binding of 125I-labelled calmodulin to this protein was demonstrated which could be antagonized with a calmodulin antagonist, trifluoperazine. These results show that the plasma membrane contains an ATP-dependent Ca2+ transporter. However, its relationship to a previously described high affinity form of Ca(2+)-stimulated Mg(2+)-dependent ATPase is not apparent because their [Mg2+] requirements to elicit maximal activity differed by two orders of magnitude.
Reinach
P S
PS
Department of Ophthalmology, Washington University School of Medicine, St. Louis, MO.
Holmberg
N
N
Chiesa
R
R
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000032:analysis*; D002147:Calmodulin / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D002352:Carrier Proteins / Q000032:analysis*; D002417:Cattle; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology*; D003315:Cornea / Q000187:drug effects / Q000201:enzymology*; D004848:Epithelium / Q000201:enzymology; D007700:Kinetics; D014268:Trifluoperazine / Q000494:pharmacology
D002147:Calmodulin; D002352:Carrier Proteins; D014268:Trifluoperazine; D000001:Calcimycin; D000255:Adenosine Triphosphate; D000252:Calcium-Transporting ATPases; D002118:Calcium