Positions of the sites labeled by N-cyclohexyl-N'-(4-dimethylamino-1-naphthyl)carbodiimide on the (Ca2+ + Mg2+)-ATPase

Biochim Biophys Acta. 1989 Feb 13;979(1):113-20. doi: 10.1016/0005-2736(89)90530-0.

Abstract

N-Cyclohexyl-N'-(4-dimethylamino-1-naphthyl)carbodiimide (NCD-4) labels (Ca2+ + Mg2+)-ATPase at Ca2+-protectable sites, believed to be at or near the two Ca2+ binding sites on the ATPase, and at nonspecific sites. The labeled ATPase has been reconstituted into lipid bilayers containing phosphatidylethanolamine labeled with fluorescein isothiocyanate. The distance between NCD-4 and fluorescein groups was measured using Forster energy transfer and the NCD-4 labels were found to be approx. 20 A from the lipid/water interface suggesting that the Ca2+ binding sites on the ATPase are also 20 A from the lipid/water interface. Addition of vanadate causes no change in the efficiency of energy transfer, suggesting that the Ca2+ binding sites on the E1 conformation of the ATPase do not move significantly with respect to the lipid/water interface in the E1-E2 transition.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding Sites
  • Ca(2+) Mg(2+)-ATPase*
  • Calcium-Transporting ATPases*
  • Carbodiimides*
  • Energy Transfer
  • Lipid Bilayers
  • Membrane Proteins / ultrastructure*
  • Phosphatidylcholines
  • Sarcoplasmic Reticulum / enzymology
  • Spectrometry, Fluorescence
  • Vanadium / pharmacology

Substances

  • Carbodiimides
  • Lipid Bilayers
  • Membrane Proteins
  • Phosphatidylcholines
  • Vanadium
  • N-cyclohexyl-N'-(4-dimethylamino-alpha-naphthyl)carbodiimide
  • Ca(2+) Mg(2+)-ATPase
  • Calcium-Transporting ATPases
  • 1,2-oleoylphosphatidylcholine