title: Mn(II)-EPR measurements of cation binding by aequorin.
abstract: Cation binding at 5 degrees C by aequorin, a bioluminescent protein from the jellyfish Aequorea victoria, was examined by means of Mn(II) EPR. The bioluminescence of aequorin is triggered by Ca(II), as well as by trivalent lanthanides, and is inhibited by Mg(II) and Mn(II). Three EF-hand Ca(II)-binding domains have been identified in the aequorin amino acid sequence. In the work reported here, active native aequorin was found to have a single tight binding site for Mn(II) with an association constant of 0.566 microM-1. Ca(II) and La(III) competed for the Mn(II) site with association constants of 1.92 microM-1 and 1.38 microM-1, respectively. The affinity of Ca(II) and La(III) for their two other (presumed) sites on aequorin was an order of magnitude less than their affinity for the Mn(II) site. Mg(II) competed for the Mn(II) site as well but with a much smaller association constant of 0.0109 microM-1. Ca(II)-independent discharged aequorin did not bind Mn(II) to a significant degree. Conjectures on the location of the Mn(II) site in the aequorin amino acid sequence and on the relationship between the binding parameters of the cations and their influence on aequorin activity are given.
authors: Department of Physics, Indiana University-Purdue University, Indianapolis 46205-2810.
authors: M D
authors: MD
authors: Kemple
authors: M L
authors: ML
authors: Lovejoy
authors: B D
authors: BD
authors: Ray
authors: F G
authors: FG
authors: Prendergast
authors: B D
authors: BD
authors: Nageswara Rao
mesh_terms: D000331:Aequorin; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D002412:Cations; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods; D007700:Kinetics; D007811:Lanthanum / Q000378:metabolism*; D008164:Luminescent Proteins; D008274:Magnesium / Q000378:metabolism*; D008345:Manganese / Q000378:metabolism*; D008433:Mathematics; D011485:Protein Binding
chemical_list: D002412:Cations; D008164:Luminescent Proteins; D008345:Manganese; D000331:Aequorin; D007811:Lanthanum; D008274:Magnesium; D002118:Calcium