The Interaction of the Lanthanide Ions with Staphylococcal Nuclease
Bruce Furie 1, Ann Eastlake 1, Alan N. Schechter 1, and Christian B. Anfinsen 1
From the
1 From the Laboratory of Chemical Biology, National Institute of Arthritis, Metabolism, and Digestive Diseases, National Institutes of Health, Bethesda, Maryland 20014
The metal ion-binding site of staphylococcal nuclease has been studied at pH 7.0 by using several lanthanide ions. The binding of Gd(III) or Nd(III) to nuclease was associated with tyrosine perturbations in the ultraviolet spectrum which were shown to include tyrosine-85 and tyrosine-115 by using mononitrotyrosine nuclease derivatives. A binding constant of 9 µm was estimated by spectrophotometric titration of nitrotyrosine-115-nuclease with Gd(III). The Ca(II) required for the binding of nuclease to deoxythymidine 3',5'-diphosphate-Sepharose could be replaced by Nd(III). Nd(III) stabilized liganded and unliganded nuclease to trypsin digestion. None of the lanthanide ions replaced Ca(II) in the generation of nuclease activity toward DNA under the conditions employed. Kinetic data compatible with a model of competitive inhibition of Ca(II) by La(III), Pr(III), Nd(III), Gd(III), Dy(III), and Yb(III) yielded inhibition constants of 1 to 2 µm. These data suggest that the lanthanide ions bind tightly to the Ca(II) binding site of nuclease in the presence or absence of substrate or substrate analogue. As does Ca(II), these ions also enhance the binding of substrate analogue to nuclease.
Submitted on April 5, 1973
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