Faculty Publications

Spectroscopic Characterization of Inhibitor Interactions With the Mn(III)/Mn(IV) Core in Lactobacillus plantarum Manganese Catalase

Document Type

Article

Publication Date

January 1997

Abstract

X-ray absorption (XAS), UV−visible, electron paramagnetic resonance (EPR), and electron spin echo envelope modulation (ESEEM) spectroscopies have been used to characterize the interaction of azide and cyanide with the Mn(III)(μ-O)2Mn(IV) site in superoxidized Mn catalase. The addition of azide causes no significant change in the X-ray absorption near edge structure (XANES) region and only minor changes in the extended X-ray absorption fine structure (EXAFS) spectra, consistent with only minimal changes in Mn−ligand geometry. In contrast, addition of either azide or cyanide causes an approximately 3-fold increase in the intensity of the visible absorption bands and a small (approximately 4%) decrease in the Mn hyperfine coupling. Anion-binding titrations indicate cooperativity in anion binding. ESEEM experiments on the azide- and cyanide-free enzyme reveal hyperfine (A = 2.88 MHz) and electric quadrupolar couplings (e2qQ = 2.29 MHz and η = 0.58) for a single 14N nucleus of the protein. These parameters are slightly altered upon addition of azide or cyanide, but ESEEM studies with 15N-labeled versions of these inhibitors show that the altered modulation is also due to protein-derived 14N. The ESEEM experiments show no evidence for coupling of azide- or cyanide-derived nitrogens to the Mn cluster. The outer shell scattering in the EXAFS suggests coordination of histidines to the binuclear Mn cluster, and the quadrupolar couplings observed for the protein-derived ESEEM detectable 14N nucleus are consistent with those expected for a histidine imidazole coordinated to Mn(IV). Taken together, these results suggest a model in which azide binds to a protein-derived site, rather than binding directly to either Mn. Despite this indirect binding, azide causes minor perturbations in the Mn2(μ-O)2 geometry, consistent with a slight flattening of the Mn2 core.

Journal Title

J. Am. Chem. Soc.

Volume

119

Issue

39

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