Jean-Claude Bunzli - Bovine Alpha-Lactalbumin - Identification of 2 Metal-Ion-Binding Sites Using the Europium(Iii) Luminescent Probe

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  Bunzli,J.C.G., Pfefferle,J.M. Helvetica Chimica Acta  1994 77 (1) 323-333

  Abstract: The luminescent Eu-III ion has been   used to probe the metal-binding sites of bovine alpha-lactalbumin   (BLA) in D2O. Upon addition of apo-BLA to an Eu-III-containing   solution, the intrinsic luminescence of the protein is quenched,   and the Eu-III luminescence is enhanced. Luminescent titrations   point to there being at least two different metal-binding sites   in the ape-protein. Curve analysis of the high resolution   D-5(0)<--F-7(0) excitation spectra reveals the existence of   three different environments for the bonded Eu-III ions. Two   environments, labelled I-a and I-b, give D-5(0)<--F-7(0) bands   very close in energy; they contain four negatively charged groups   and are assigned to one site we identify as the calcium-binding   site. Site I is protected from solvent influences and is somewhat   rigid, since it displays selectivity towards lanthanide ions. The   origin of the two similar environments I-a and I-b could not be   determined unambiguously. The third environment is ascribed to a   nonspecific metal-binding site in which the Eu-III ion is more   exposed to the solvent (site II). It is sequentially populated   after saturation of site I, and its population is pH-dependent.   The affinity constant of Eu-III for this site was estimated from   the excitation spectra: log K-2(app) = 3.5(1). Assignment of the   metal binding sites has been facilitated by comparison with model   compounds, [Eu(dota)](-) (dota = 1,4,7,10-tetraazacyclododecane   N,N',N'',N'''-tetraacetate), [Eu(dtpa)](2-) (dtpa =   diethylenetriamine tetraacetate), and [Eu(bsa)] (bsa = bovine   serum albumin). The usefulness and limits of the use of   curve-analysis procedures to unravel the various components of   D-5(0)<--F-7(0) excitation spectra in biological materials are   also discussed

  Address (URL): WOS:A1994MW15900030