MCD-EPR studies of deoxy[FeII,FeII]hemerythrin: probes of endogenous bridging ligands and exogenous ligand binding

A quantitative interpretation is presented for EPR spectra from integer-spin metal centers having large zero-field splittings. Integer-spin, or non-Kramers, centers are common in metalloproteins and many give EPR signals, but a quantitative understanding has been lacking until now. Heterogeneity of the metal's local environment will result in a significant spread in zero-field splittings and in broadened EPR signals. Using the spin Hamiltonian Hs = S.D.S + beta S.g.B and some simple assumptions about the nature of the zero-field parameter distributions, a lineshape model was devised which allows accurate simulation of single crystal and frozen solution spectra. The model was tested on single crystals of magnetically dilute ferrous fluosilicate. Data and analyses from proteins and active-site models are presented with the microwave field B1 either parallel or perpendicular to B. Quantitative agreement of observed and predicted signal intensities is found for the two B1 orientations. Methods of spin quantitation are given and are shown to predict an unknown concentration relative to a standard with known concentration. The fact that the standard may be either a non-Kramers or a Kramers center is further proof of the model's validity. The magnitude of the splitting in zero magnetic field is of critical importance; it affects not only the chance of signal observation, but also the quantitation accuracy. Experiments taken at microwave frequencies of 9 and 35 GHz demonstrate the need for high-frequency data as only a fraction of the molecules give signals at 9 GHz.

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“…EPR signals have been reported from the binuclear ferrous site of the azide complex of hemerythrin (3,24). Fig.…”

Section: Azide Complex Of Hemerythrinmentioning

confidence: 99%

Integer-spin electron paramagnetic resonance of iron proteins

Hendrich¹,

Debrunner²

1989

Biophysical Journal

216

190

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“…51 shows the EPR spectra obtained for ligands representative of both groups.Within the first group of ligands, azide is unique in that in addi tion to bleaching the deoxyNO color, the product exhibits an EPR signal with g-value at 12,3. Such a signal was previously observed for deoxyNg"(6). Thus, In the case of azide, cyanate, and fluoride, the colorless products are ligand adducts of deoxyHr.…”

supporting

confidence: 72%

“…Table 3,11, Resonance Raman vibrational data for semi-metNO Table 3.12, Extinction coefficients for deoxyNO 191 Table 3.13, Double integrations of the EPR signal for deoxyNO 197 Table 3,14, Experimental determinations of a for deoxyNO Table 3,18, Estimated extinction coefficients for deoxyF"NO Table 3,19, Double integrations of the EPR signal for deoxyF"NO Table 3,20. Resonance Raman data for deoxyF"NO at 90 K using 647,1 nm excitation Table 3,21, Equilibrium constants for the binding of nitric oxide to deoxyHr Table 3,22, Double integration of the EPR signal for deoxyNO as a function of the incubation period for sulfide Table 4,3, Affinity constants (M"i) for the binding of XO (X = N, 0, C) to Hb, Mb, and Hr 290 ferrous atoms mediated by a hydroxo bridge (6,7).…”

Section: Reduction Potentials In Volts For Hemerythrin Derivativesmentioning

confidence: 99%

The reactions of nitric oxide and nitrite with hemerythrin

Nocek

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“…The O z binding site of deoxyhemerythrin consists of two high-spin ferrous atoms that are bridged by a hydroxo and two carboxylato groups; one coordination site is available for exogenous ligand binding. The addition of azide to deoxyhemerythrin results in the formation of an aquo bridge (30) and a ferromagnetic interaction between the irons of 3 cm" 1 (31). In this state EPR signals have been observed at 9 and 34 GHz as shown in Fig.…”

Section: Iron-oxo Clustersmentioning

confidence: 87%