Absorption spectra of sperm-whale ferrimyoglobin

1. Sperm-whale ferrimyoglobin was found to contain 0.308% of Fe, on a dry weight basis, corresponding to a molecular weight of 18130. The solid takes up moisture to an equilibrium state, and was then assayed to contain 0.280% of Fe. 2. Absorption spectra are presented for acidic ferrimyoglobin, Fe(+)(H(2)O), and its conjugate base, Fe-OH, as well as for the fluoride and cyanide complexes, within the range 200-2500mmu. Data for ferromyoglobin-carbon monoxide, Fe(II)-CO, in the visible region are also included. 3. Minor spectral differences have been found between whale and horse myoglobins, particularly in the effect of temperature on the visible-absorption spectrum of Fe-OH.     

Analysis of the visible spectra of some sperm-whale ferrimyoglobin derivatives

The visible spectra of sperm-whale metmyoglobin and its fluoride, hydroxide, cyanide, azide, cyanate, formate, acetate, thiocyanate and nitrite derivatives were measured and resolved into Gaussian components. A linear correlation between the intensity of the band at 18500cm.⁻¹ and the magnetic susceptibility is found, suggesting that the band is the β-band of the low-spin form. The analysed intensity of this band can be used to determine the relative proportions of the two spin states present. The spectra of the pure high-spin and low-spin components of each derivative can be reconstructed. Correlations between the energy of the near-infrared band and the visible bands of the high-spin form, and between the position and intensity of the 16000cm.⁻¹ band, are demonstrated. The relationship between the position of the Soret band and the magnetic susceptibility can be explained in terms of the equilibrium between spin states.     

A proton magnetic relaxation study of ferrimyoglobin in aqueous ionic solutions

Proton magnetic longitudinal T₁ relaxation times have been measured for acid (horse) ferrimyoglobin solutions [0.1 M NaCl and KH₂PO₄, 2 M NaCl and 1 M MgCl₂] from 5°C to 35°C in dependence on myoglobin concentration up to 6 mM. The enhancement of the relaxation rate due to the paramagnetic haem iron. which is observed in this temperature range is compared with analogous data for the ferrihaemoglobin solution. The conclusion is that the protons exchanging from the haem pocket with bulk solvent are not those from the water molecule at the sixth ligand site of haem iron. The exchanging protons are more than 4 Å away from the haem iron being closer to it in ferrimyoglobin than in ferrihaemogiobin. This distance becomes larger in solutions with higher salt concentration, the largest difference between 0.1 M NaCl and 1 M MgCl₂ being over one Angstrom unit. This indicates a conformational change of the haem pocket, possibly its tightening.     

Analysis of the principal g-tensors in single crystals of ferrimyoglobin complexes

This paper describes a method of determining the directions of the principal axes of the g-tensors in single crystals from measurements of the g-value variation in three crystalline planes (ab, bc*, ac) and of the principal values of g-tensors. Measurements of paramagnetic resonance spectra of ferrimyoglobin (Mb(Fe3+)) complexes (Mb(Fe3+).CN-, .N3-, .imidazole(Im.) and .OCN-) in single crystals provided detailed information on the electronic state of the Fe3+. The direction of the z axis in Mb(Fe3+).CN-, .N3- and .Im. is not parallel to that in Mb(Fe3+).H2O, which has been used as the haem normal, where the z axis is one of the principal axes of the g-tensors corresponding to the maximal (in low-spin state of Fe3+ in haem) or minimal (in high-spin state of Fe3+ in haem) g-value. In Mb(Fe3+).OCN-, however, which is in thermal equilibrium between high-spin and low-spin states, the directions of the z axis in both states seem to be perpendicular to the haem plane. The direction of the x axis is not parallel to the plane of the linked histidine ring.     

Proton hyperfine resonance assignments in trimethylphosphine ligated ferrimyoglobin using saturation transfer spectroscopy

1H-NMR saturation transfer spectroscopy and nuclear Overhauser effect (NOE) have been utilized to assign several heme resonances in the low-spin trimethyl phosphine complex of sperm whale metmyoglobin. The two methods permit the location of the heme methyl resonances and the heme 2-vinyl group resonances. A qualitative comparison involving the methyl shift pattern in metMbN3, metMbCN, imidazole metMb and trimethyl phosphine metMb shows a reverse methyl shift between pyrrole I and pyrrole IV. The different hyperfine shift pattern for metMbPMe3 is suggested to arise from: (i) a possible reorientation of the proximal histidine plane; (ii) different heme protein contacts in the different ligated proteins, and (iii) a small contribution from high-spin character. The 2-vinyl group is formed in the cis plane orientation.     

Solvent accessibility calculations for sperm whale ferrimyoglobin based on refined crystallographic data

The calculation of solvent accessibility parameters from protein crystallographic data, by the method of Lee and Richards has been simplified to allow treatment of single atoms and their immediate environment. New accessibility values for all titratable amino acid residues in myoglobin have been computed from recent X-ray structure data of Takano. A number of prominent differences appear between these values and those from the older structure data of Watson. Differences are interpreted in terms of the proximity of neighboring residues.     

Formation of a steady state in the radiolysis of ferrimyoglobin in aqueous solution

The interaction of radiation-generated · OH radicals with ferrimyoglobin in deaerated aqueous solution at neutral pH has been quantitatively studied. Changes in the visible absorption spectrum have been analyzed on the basis of composition changes of the ferri, deoxy, and ferriperoxide forms of the metalloprotein. A postirradiation thermal process must be considered in order to evaluate the radical-induced composition changes. Initially, ·OH induces reduction of ferrimyoglobin to the deoxy form with a G value (molecular yield/100 eV of absorbed energy) in the zero-dose limit of 1.4 (±0.2). Radiation-generated H₂O₂ reacts with the ferrimyoglobin substrate to produce ferrimyoglobin peroxide with a G value of 0.7 (±0.1) in the zero-dose limit. At doses of >1 krad μm^?1 of myoglobin present, the composition of the three myoglobin derivatives reaches a radiolysis steady state. In this moderate-dose plateau region, this composition is 44% ferri, 18% deoxy, and 38% ferri peroxide. The · OH-induced hemoprotein radicals that do not initiate 1-eq redox conversions undergo reactions that generate dimer and other globin-modified material.