Dynamics and structure in the Mn2+ site of concanavalin A as determined by high-field EPR and ENDOR spectroscopy

Abstract

The properties of the Mn2+ site in the protein concanavalin A were investigated by single crystal W-band EPR/ENDOR (electron-nuclear double resonance) measurements. Initially, room temperature EPR measurements were carried out, one type of Mn2+ was identified and its zero-field splitting (ZFS) tensor was determined. In contrast, low temperature EPR measurements showed that two chemically inequivalent Mn2+ are present, Mn(A)2+ and Mn(B)2+, differing in their ZFS tensors. Variable temperature measurements revealed a two-site exchange between the two types. Although the dynamic process has been characterized by its rate and activation energy, just from the EPR measurements it was not possible to assign it to a specific residue. 1H ENDOR measurements of the water and imidazole protons, which are the main contributors to the ENDOR spectra, showed only one type of signals, namely, they were not sensitive to the differences between Mn(A)2+ and Mn(B)2+. 55Mn ENDOR spectra, which are dominated by the 55Mn isotropic hyperfine, a(iso), and the nuclear quadrupole interaction did sense the differences. Analysis of the spectra recorded with the magnetic field along the crystallographic axes showed that the two have the same a(iso) but different quadrupole tensors.

Effect of Ca2+/Sr2+ Substitution on the Electronic Structure of the Oxygen-Evolving Complex of Photosystem II: A Combined Multifrequency EPR, 55Mn-ENDOR, and DFT Study of the S2 State

“…The EPR settings are the same as described in the main text. Mn 2+ has two characteristic peaks located the RF of ~114 MHz, ~158 MHz and at 375 MHz (not shown) under these experimental conditions described in Figure caption 2, see also ref[6][7][8] .…”

Structure and Nature of Manganese(II) Imidazole Complexes in Frozen Aqueous Solutions

“…More precisely, there was negligible distribution in the proton hyperfine interactions, in particular the dipolar couplings, which are determined by the Mn–H distances. Studies on single crystals of ConA have shown that the manganese ions occupy two sites that have different zero-field interactions ( D = 789 and E = 189 MHz and D = 970 and E = 140 MHz) but the same proton ENDOR spectra, as well as 55 Mn quadrupole tensors. , This suggested that the Mn II zero-field interactions are more sensitive than the other magnetic interactions to local structure disorder or, more likely, to nonstructural effects. Electrostatics originating from well beyond the primary ligand sphere have been shown to affect the Mn II zero-field interaction with little or no change to the structure of the ligand sphere. , Therefore, it seemed likely that large zero-field distribution of the imidazole complexes originated from the disorder in the surrounding ions that did not affect the structure of the manganese imidazole complex.…”

pH-Dependent Structures of the Manganese Binding Sites in Oxalate Decarboxylase as Revealed by High-Field Electron Paramagnetic Resonance

“…However, conventional EPR has proven to be of limited value due to poor resolution and high spectral complexity. Recently, several experiments have been carried out using high magnetic-field high-frequency EPR (HFEPR) − that suggest that this approach not only circumvents these two limitations but also allows accurate measurement of the magnetic spin parameters of an Mn(II) ion from which valuable information about its electronic structure and interactions with the environment can be extracted.…”