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Biomedical subjects

M J Barber

Publications and source records attributed to M J Barber.

At least 109 records · Page 6Linked to original sources
PubMed1983

Formate dehydrogenase from Methanobacterium formicicum. Electron paramagnetic resonance spectroscopy of the molybdenum and iron-sulfur centers.

Formate dehydrogenase from Methanobacterium formicicum was examined by electron paramagnetic resonance spectroscopy. Although oxidized enzyme yielded no EPR signals over the temperature range 8-200 K, dithionite reduction resulted in generation of two paramagnetic components. The first, a nearly isotropic signal visible at temperatures below 200 K with g1 = 2.018, g2 = 2.003, and g3 = 1.994, exhibited nuclear hyperfine interaction with two equivalent protons (A1 = 0.45, A2 = 0.6, and A3 = 0.55 milliTeslas). EPR spectra of partially reduced 95Mo-enriched formate dehydrogenase exhibited additional 3-4 milliTeslas splittings, due to spin interaction with the 95Mo nucleus. Thus, this signal is due to a Mo center. This is the first reported example of a Mo center with gav greater than 2.0 in a biological system. The second species, a rhombic signal visible below 40 K with g values of g1 = 2.0465, g2 = 1.9482, and g3 = 1.9111 showed no hyperfine coupling and was assigned to reduced Fe/S. Both paramagnetic species could be detected in samples of M. formicicum whole cells anaerobically reduced with sodium formate. The Mo(V) signal was altered following addition of cyanide (g1 = 1.996, g2 = 1.988, and g3 = 1.980). Growth of bacteria in the presence of 1 mM WO4(2-) resulted in abolition of the Mo(V) EPR signal and formate dehydrogenase activity. Em, 7.7 was -330 mV for Mo(VI)/Mo(V) and -470 mV for Mo(V)/Mo(IV).

Aldehyde Oxidoreductases↗
PubMed1983

Studies of the mobility of maleimide spin labels within the erythrocyte membrane.

We have confirmed a method yielding reproducible and reliable spectrometric parameters derived from spin-labeled erythrocyte ghosts using nitroxide derivatives of maleimide compounds. The disorder parameter, W/S, was shown to vary with changes in the structure of the label, the conditions utilized for labeling such as ionic strength and erythrocyte age and the presence of drugs such as alcohol and acetaminophen. The nitroxide spectrum was also found to change with increasing and decreasing temperature in an irreversible manner. These findings should permit increased reliance to be placed on the spin-labeling technique when used to monitor changes in membrane lipid or protein assembly.

Cyclic N-Oxides↗
PubMed1983

Disruption of erythrocyte membranal organization by superoxide.

Human erythrocyte ghosts were covalently labeled with 4-maleimide-2,2,6,6-tetramethylpiperidinooxyl. Electron paramagnetic resonance (EPR) spectrometry revealed two major binding environments representing strongly (S) and weakly (W) immobilized species. The disorder parameter, W/S, determined from the respective peak amplitudes, was shown to be irreversibly elevated following treatment of the labeled ghosts with superoxide, indicating an increase in membrane fluidity. Labeled ghosts reduced with ascorbate showed no nitroxide EPR signals. However, following exposure of these membranes to superoxide, the nitroxide spectrum returned with a W/S ratio of 25. In contrast, the disorder parameter for spin-labeled ghosts decreased following exposure to hydroxyl radicals suggesting decreased fluidity, as a result of lipid peroxidation. This effect could be prevented by the inclusion of mannitol. These changes in membrane fluidity and/or protein mobility observed by EPR are compared with previous results obtained by other methods and provide additional evidence for physiologic alterations initiated by superoxide.

Cyclic N-Oxides↗
PubMed1983

Influence of the autonomic nervous system on the genesis of cardiac arrhythmias.

The purpose of this report is to present a brief overview of the concepts concerned with the potential effects of the autonomic nervous system on the genesis of cardiac arrhythmias. First, we present data indicating the functional importance of presynaptic and postsynaptic vagal modulation of adrenergic neurotransmission on electrical activity in sinus and A-V nodes, and contractile activity of canine cardiac Purkinje fibers. Second, we indicate the possible intraventricular adrenergic and vagal pathways to the ventricles, suggesting that sympathetic fibers travel in the epicardium while vagal fibers appear to cross the A-V groove in the epicardium but then take an intramural route for part of their course. Finally, we demonstrate that a lesion such as a myocardial infarction can interrupt neural fibers traveling through the infarction to affect noninfarcted myocardium distal to the infarction. These factors may play a role in the genesis of some cardiac arrhythmias.

Adrenergic Fibers↗
PubMed1983

Evidence for formation of superoxide and formate radicals in Methanobacterium formicicum.

Using spin labeling and spin trapping techniques in combination with electron paramagnetic resonance spectrometry, we have detected the formation of superoxide by whole cells of Methanobacterium formicicum under aerobic conditions in the presence and absence of sodium formate. Rates of superoxide generation have been estimated. The formation of additional free radical species, including formate, was observed. Production of these and other free radicals resulted in lipid peroxidation and concomitant cell damage.

Electron Spin Resonance Spectroscopy↗
PubMed1983

Selective denervation of the heart.

Total denervation of the canine heart consisted of intrapericardial neural dissection of the left atrium, left superior pulmonary vein, and main pulmonary artery and cutting of the ventrolateral cardiac nerve (stage I). The fat pad and all nerves were removed from between the pulmonary artery and aorta (stage II). Dissection proceeded from the pericardial reflection along the superior vena cava to the azygos vein, which was cleared, double tied, and cut. The right pulmonary artery was cleaned, and the superior right atrium was dissected to its intersection with the left atrium (stage III). Denervation was tested by electrical stimulation of both vagi and stellate ganglia, while recording inotropic, chronotropic, and dromotropic events, before and after each stage. Stage I deleted most left autonomic input to the heart without interrupting right sympathetics. Stage II completed left autonomic denervation but preserved much of the right sympathetic input. Large nerves along the dorsal surface of the pulmonary artery carried inputs from both left and right sympathetics. Stage III completed the denervation of atrioventricular and sinoatrial nodal structures and removed all remaining ventricular inotropic influences. Selective denervation of atrioventricular and sinoatrial nodal regions appears feasible for preparation of chronic canine models.

Animals↗
PubMed1982

Properties of the prosthetic groups of rabbit liver aldehyde oxidase: a comparison of molybdenum hydroxylase enzymes.

Rabbit liver aldehyde oxidase (AO), like milk xanthine oxidase (XO) and chicken liver xanthine dehydrogenase (XDH), possesses the following prosthetic groups: FAD, a functional Mo center, and two spectroscopically distinct iron-sulfur centers, one with gav less than 2.0 (termed Fe/S I) and the other with gav greater than 2.0 (termed Fe/S II) in the reduced enzyme. EPR spectra for the Mov species were found to be nearly identical in AO and XO for a number of enzyme complexes, and the midpoint reduction potentials for functional MoVI/MoV (-359 mV) and MoV/MoVI (-351 mV) were nearly the same in all three enzymes (50 mM phosphate, pH 7.8). A strong magnetic interaction between MoV and reduced Fe/S I, previously detected in XO and XDH, was also found in AO. No MoV-Fe/S II interaction could be detected in AO (nor in XO). In contrast, the order of reduction of Fe/S I and Fe/S II, as measured from their midpoint potentials, is reversed in AO (Em = -207 and -310 mV, respectively) as compared to XO (Em = -280 and -245 mV, respectively) in phosphate buffer at pH 7.8. The oxidized-reduced extinction coefficients at 450 and 550 nm for the two centers are also apparently reversed in AO and XO. Although magnetic interaction between FAD and one or both reduced Fe/S centers has been detected in both AO and XO, no magnetic interaction between the two reduced Fe/S centers themselves was found in AO (although such interaction has been seen in XO). The average FAD reduction potential is substantially more positive in AO (Em for FAD/FADH., -258 mV; FADH./FADH2, -212 mV at pH 7.8) than in XO or XDH. It can be concluded that although the properties and immediate environment of the functional Mo center are conserved in the three Mo hydroxylase enzymes, and all three enzymes possess the same set of prosthetic groups, the properties of the groups which transfer electrons from the Mo to the ultimate electron acceptor can vary substantially in AO, XO, and XDH.

Aldehyde Oxidase↗
PubMed1982

Escherichia coli sulfite reductase hemoprotein subunit. Prosthetic groups, catalytic parameters, and ligand complexes.

Escherichia coli NADPH-sulfite reductase can be dissociated into an oligomeric flavoprotein and a monomeric hemoprotein (HP) subunit in 4 M urea. HP catalyzes stoichiometric 6-electron reductions of SO32- (to S2-) and of NO2-, as well as 2-electron reduction of NH2OH, with reduced methyl viologen (MV+) as reductant. While Vmax values are highest with the nitrogenous substrates, Km for SO32- is 2 to 3 orders of magnitude less than the Km for NO2- or NH2OH. EPR spectroscopic and chemical analyses show that HP contains one siroheme and one Fe4S4 center per polypeptide. The heme is in the high spin Fe3+ state in HP as isolated. Near-quantitative reduction of the Fe4S4 center to a state yielding a g = 1.94 type of EPR spectrum by S2O42- and/or MV+ could be achieved if HP was converted to either the CN- or CO complex or treated with 80% dimethyl sulfoxide. HP binds one SO32- or CN- per peptide. Binding of these ligands, as well as CO, appears to be mutually exclusive and to involve the heme. The heme Fe3+/Fe2+ potential is shifted from -340 mV in the free HP to -155 mV in the HP-CN- complex. The potential of the Fe4S4 center is approximately 70 mV more negative in the CN- as opposed to the CO-ligated HP (-420 mV), a result which indicates the presence of heme-Fe4S4-ligand interaction in the HP complexes.

Cyanides↗
PubMed1982

Magnetic interactions in milk xanthine oxidase.

The relaxation behavior of the EPR signals of MoV, FAD semiquinone, and the reduced Fe/S I center was measured in the presence and absence of other paramagnetic centers in milk xanthine oxidase. Specific pairs of prosthetic groups were rendered paramagnetic by poising the native enzyme or its desulfo glycol inhibited derivative at appropriate potentials and pH values. Magnetic interactions were found between the following species: Mo--Fe/S I (100-fold increase in microwave power required to saturate the MoV EPR signal at 103 K when Fe/S I is reduced as opposed to oxidized), FAD--Fe/S I and FAD--Fe/S II (70-fold increase in power required to saturate the FADH.EPR signal at 173 K when either Fe/S center is reduced), and Fe/S I--Fe/S II (2.5-fold increase in power to saturate the reduced Fe/S I EPR signal at 20 K when Fe/S II is reduced). The Mo--Fe/S I interaction was also detected as a reduced Fe/S I induced splitting of the MoV EPR spectrum at 30 K. No splittings of the FADH. or Fe/S center spectra were detected. No magnetic interactions were found between FAD and Mo or between Mo and Fe/S II. These results, together with those of Coffman & Buettner [Coffman, R. E., & Buettner, G. R. (1979) J. Phys. Chem. 83, 2392-2400], were used to estimate the following approximate distances between the electron carrying prosthetic groups of milk xamthine oxidase: Mo--Fe/S I, 11 +/- 3 A; Fe/S I-Fe/S II, 15 +/- 4 A; FAD-Fe/S I, 16 +/- 4 A; FAD-Fe/S II, 16 +/- 4 A. A model for the arrangement of these groups within the xanthine oxidase molecule is suggested.

Animals↗