Intrapericardial denervation of the heart.
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Animals↗
Biomedical subjects
M J Barber
Publications and source records attributed to M J Barber.
Electron-paramagnetic-resonance spectroscopy of complexes of xanthine oxidase with xanthine and uric acid.
Molybdenum(V) e.p.r. signals from reduced functional milk xanthine oxidase molecules (the Rapid signals), obtained in the presence of purine substrates and products, were further investigated [cf. Bray & Vänngård, (1969) Biochem. J. 114, 725-734; Pick & Bray (1969) Biochem. J. 114, 735-742]. Xanthine forms two complexes with the enzyme that are believed to correspond to different orientations of the substrate molecule in the active site. Only one complex appears to undergo the catalytic reaction. Non-productive complexes, analogous to theone with xanthine, are not formed by 1-methylxanthine or purine. Uric acid forms more than one e.p.r.-detectable complex, one of which is analogous to the non-productive xanthine complex. The computer program used for handing the e.p.r. data is described briefly.
Animals↗
Changes in apparent pH on freezing aqueous buffer solutions and their relevance to biochemical electron-paramagnetic-resonance spectroscopy.
Changes in apparent pH occurring during fast freezing of aqueous buffer solutions and cooling to -196 degrees C were studied by various semiquantitative methods, including simple visual measurements of colour changes with pH indicators, as well as measurements of pH-dependent changes in the e.p.r. (electron paramagnetic resonance) spectra of solutions of three different metalloenzymes. It is concluded that apparent pH changes of up to about 3pH units may occur under particular conditions. Such changes were independent of the time taken to freeze the samples, when this was varied from about 3ms t0 20s, but were affected by the presence of some proteins in solution. Recommendations on the buffers that should be used to avoid such apparent pH changes in e.p.r. spectroscopy and other low-temperature biochemical work are made. Phosphate and pyrophosphate buffers, which gave large decreases (2-3 pH units), and Tris, which under some conditions gave increases of about the same magnitude, are to be avoided. Certain zwitterionic buffers such as Bicine [NN-bis-(2-hydroxyethyl)glycine] are satisfactory. Apparent pH effects were found to depend on buffer and protein concentration. It is therefore recommended that as a prelude to future detailed low-temperature biochemical work, appropriate tests with an indicator system should be performed.
Buffers↗
Oxidation--reduction potentials of turkey liver xanthine dehydrogenase and the origins of oxidase and dehydrogenase behaviour in molybdenum-containing hydroxylases.
Redox potentials for the various centres in the enzyme xanthine dehydrogenase (EC 1.2.1.37) from turkey liver determined by potentiometric titration in the presence of mediator dyes, with low-temperature electron-paramagnetic-resonance spectroscopy. Values at 25 degrees C in pyrophosphate buffer, pH 8.2, are: Mo(VI)/Mo(V)(Rapid),-350 +/- 20mV; Mo(V) (Rapid)/Mo(IV), -362 +/- 20mV; Fe-S Iox./Fe-S Ired., -295 +/- 15mV; Fe-S IIox./Fe-S IIred., -292 +/- 15mV; FAD/FADH,-359+-20mV; FADH/FADH2, -366 +/- 20mV. This value of the FADH/FADH2 potential, which is 130mV lower than the corresponding one for milk xanthine oxidase [Cammack, Barber & Bray (1976) Biochem. J. 157, 469-478], accounts for many of the differences between the two enzymes. When allowance is made for some interference by desulpho enzyme, then differences in the enzymes' behaviour in titration with xanthine [Barber, Bray, Lowe & Coughlan (1976) Biochem. J. 153, 297-307] are accounted for by the potentials. Increases in the molybdenum potentials of the enzymes caused by the binding of uric acid are discussed. Though the potential of uric acid/xanthine (-440mV) is favourable for full reduction of the dehydrogenase, nevertheless, during turnover, for kinetic reasons, only FADH and very little FADH2 is produced from it. Since only FADH2 is expected to react with O2, lack of oxidase activity by the dehydrogenase is explained. Reactivity of the two enzymes with NAD+ as electron acceptor is discussed in relation to the potentials.
Animals↗
Oxidation-reduction potentials of molybdenum, flavin and iron-sulphur centres in milk xanthine oxidase.
1. The mid-point reduction potentials of the various groups in xanthine oxidase from bovine milk were determined by potentiometric titration with dithionite in the presence of dye mediators, removing samples for quantification of the reduced species by e.p.r. (electron-paramagnetic-resonance) spectroscopy. The values obtained for the functional enzyme in pyrophosphate buffer, pH8.2, are: Fe/S centre I, -343 +/- 15mV; Fe/S II, -303 +/- 15mV; FAD/FADH-; -351 +/- 20mV; FADH/FADH2, -236 +/-mV; Mo(VI)/Mo(V) (Rapid), -355 +/- 20mV; Mo(V) (Rapid)/Mo(IV), -355 +/- 20mV. 2. Behaviour of the functional enzyme is essentially ideal in Tris but less so in pyrophosphate. In Tris, the potential for Mo(VI)/Mo(V) (Rapid) is lowered relative to that in pyrophosphate, but the potential for Fe/S II is raised. The influence of buffer on the potentials was investigated by partial-reduction experiments with six other buffers. 3. Conversion of the enzyme with cyanide into the non-functional form, which gives the Slow molybdenum signal, or alkylation of FAD, has little effect on the mid-point potentials of the other centres. The potentials associated with the Slow signal are: Mo(VI)/Mo(V) (Slow), -440 +/- 25mV; Mo(V) (Slow)/Mo(IV), -480 +/- 25 mV. This signal exhibits very sluggish equilibration with the mediator system. 4. The deviations from ideal behaviour are discussed in terms of possible binding of buffer ions or anti-co-operative interactions amongst the redox centres.
Aldehydes↗
A new non-functional form of milk xanthine oxidase containing stable quinquivalent molybdenum.
A new non-functional modified form of milk xanthine oxidase is described. This contains molybdenum in a quinquivalent state, which is resistant to both oxidation and reduction. The new species is derived from the native enzyme in a two-step process. The first step is the conversion into the desulpho form, via loss of the 'persulphide' sulphur, and the second involves reaction with ethylene glycol or other reagents. The species gives a characteristic Mo(V) electron-paramagnetic-resonance signal, without proton splittings, designated Resting II. This is virtually identical with signals reported previously from resting turkey liver xanthine dehydrogenase and rabbit liver aldehyde oxidase. The possibility is discussed that species Resting II, prepared with ethylene glycol, contains a -COCH2OH residue bound to a nitrogen ligand of molybdenum.
Aldehydes↗
Studies by electron-paramagnetic-resonance spectroscopy and stopped-flow spectrophotometry on the mechanism of action of turkey liver xanthine dehydrogenase.
Studies by e.p.r. (electron-paramagnetic-resonance) spectroscopy and by stopped-flow spectrophotometry on turkey liver xanthine dehydrogenase revealed strong similarities to as well as important differences from the Veillonella alcalescens xanthine dehydrogenase and milk xanthine oxidase. The turkey enzyme is contaminated by up to three non-functional forms, giving molybdenum e.p.r. signals designated Resting I, Resting II and Slow. Slow and to a lesser extent Resting I signals are like those from the Veillonella enzyme, whereas Resting II is very like a resting signal described by K. V. Rajagopolan, P. Handler, G. Palmer & H. Beinert (1968) (J. Biol. Chem. 243, 3784-3796) for aldehyde oxidase. Another non-functional form that gives the Inhibited signal is produced on treatment of the enzyme with formaldehyde. Stopped-flow measurements at 450 nm show that, as for the milk enzyme, reduction by xanthine is rate-limiting in enzyme turnover. The active enzyme gives rise to Very Rapid and Rapid molybdenum(V) e.p.r. signals, as well as to an FADH signal. That these signals are almost indistinguishable from those of the milk enzyme, confirms the similarities between the active sites. There are two types of iron-sulphur centres that give signals like those in the milk enzyme, though with slightly different parameters. Quantitative reduction titration of the functional enzyme with xanthine revealed two important differences between the turkey and the milk enzymes. First, the turkey enzyme FADH/FADH2 system has a redox potential sufficiently low that xanthine is incapable of reducing the flavin completely. This finding presumably explains the very low oxidase activity. Secondly, whereas the Fe/S II chromophore in the milk enzyme has a relatively high redox potential, for the turkey enzyme the value of this potential is lower and similar to that of its Fe/S I chromophore.
Animals↗
Iron-sulphur systems in some isolated multi-component oxidative enzymes.
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Aldehyde Oxidoreductases↗
Distribution of reducing equivalents on xanthine oxidase molecules and the rates of the intramolecular electron-transfer reactions.
When xanthine oxidase turns over 1-methylxanthine aerobically at pH8.2, the time-sequence in development of its electron-paramagnetic-resonance signals is not primarily due to slow intramolecular reactions among its centres. It derives instead from gross differences in electron distribution within enzyme molecules reduced by the substrate in comparison with those that have subsequently been partly reoxidized by one-electron reaction with oxygen.
Chemical Phenomena↗
Outcomes for patients with stroke in managed care vs fee-for-service.
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Cerebrovascular Disorders↗
The influence of halothane and nitroprusside on canine spinal cord hemodynamics.
Regional spinal cord blood flow was measured in 12 halothane-anesthetized mongrel dogs by means of 15 +/- 3 u radiolabeled microspheres under (1) control conditions, (2) 60-torr mean arterial blood pressure (MAP) using sodium nitroprusside (NPS), (3) 50-torr MAP using NPS, and (4) after reestablishment of normotension. With the onset of either level of hypotension, there were nonsignificant decreases from control conditions in cardiac output and pulmonary capillary wedge pressure (PCWP). A rise in heart rate was seen at 50 torr MAP. There were no significant changes in spinal cord blood flow (SCBF) under either hypotensive condition in any region of the cord, indicating an intact autoregulatory response. Sensitivity of SCBF to changes in the PaCO2 was shown to be 1.02 ml/min/100 g tissue per torr PaCO2. These data showed that (1) deliberate hypotension with NPS did not change SCBF in the dog; (2) sensitivity to PaCO2 was high under halothane anesthesia; and (3) SCBF was relatively homogeneous throughout the length of the cord.
Analysis of Variance↗