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Biomedical subjects

J L Johnson

Publications and source records attributed to J L Johnson.

18 recordsLinked to original sources
PubMed1979

The oxidation of sulphite in animals systems.

In animals the terminal step in the pathway for degradation of sulphur-containing amino acids is the oxidation of sulphite to sulphate. This reaction is catalysed by the enzyme sulphite oxidase. The enzyme contains molybdenum and a cytochrome b5 type haem, is localized in the mitochondrial intermembrane space and transfers electrons from sulphite to cytochrome c on the inner membrane. The sulphite oxidase protein has a molecular weight of 110 000 (chicken) to 122 000 (human) and exists as a dimer of identical subunits. The haem and molybdenum cofactors are present on separate domains of the molecule. The structure of the molydbenum cofactor has not been worked out in detail, but this cofactor is known to be present in many other molybdoenzymes including xanthine oxidase and nitrate reductase. Three cases of genetic sulphite oxidase deficiency in humans have been reported. The three affected children displayed mental retardation, neurological abnormalities and dislocated ocular lenses. The biochemical basis for lack of enzyme activity in each case has been studied. All three have been shown to lack the sulphite oxidase protein, but in one case this appears to be secondary to a defect in synthesis of the molybdenum cofactor. Sulphite oxidase deficiency has been produced in the rat by administration of high levels of tungsten. Sulphite oxidase-deficient animals are particularly susceptible to the toxic effects of sulphite and atmospheric sulphur dioxide.

Animals↗
PubMed1978

The interaction of arsenite with the molybdenum center of chicken liver xanthine dehydrogenase.

Inactivation of chicken liver xanthine dehydrogenase by arsenite is reflected in the molybdenum electron paramagnetic resonance signal at g = 1.97. The arsenite spectrum shows additional splittings and considerable broadening yet remains comparable to the native in total intensity. Further subtle alterations of the molybdenum signal of arsenite-treated enzyme are seen in the presence of purine-type substrates or inhibitors.

Animals↗
PubMed1977

Sulfite oxidase deficiency. Biochemical and clinical investigations of a hereditary metabolic disorder in sulfur metabolism.

Study of a 4 1/2-year-old boy with the unusual combination of acute infantile hemiplegia, ectopia lentis and the absence of homocystinuria showed large amounts of abnormal sulfur-containing metabolites (sulfite, thiosulfate and S-sulfocysteine) in the urine. Sulfite and S-sulfocysteine were also present in the plasma. His inorganic sulfate excretion was only 50 per cent of total sulfur, as compared with 75 to 95 per cent by controls. Loading with L-cysteine hydrochloride and L-methionine further increased the excretion of sulfite and thiosulfate, but not inorganic sulfate excretion. Sulfite oxidase activity in skin fibroblasts average 1.07 nmol of cytochrome d reduced per milligram of protein per minute in control lines; it was not detectable (less than 5 per cent) in the patient. Activity was reduced in both parents (0.50 in the father and 0.32 in the mother)--compatible with autosomal recessive inheritance. Good biochemical responses to a low sulfur amino acid diet suggest that early treatment may benefit the patient.

Amino Acids, Sulfur↗
PubMed1977

In vitro reconstitution of demolybdosulfite oxidase by molybdate.

Reconstitution of purified demolybdosulfite oxidase from rat liver has been achieved using inorganic molybdate as the source of molybdenum. The activation process has a pH optimum of 7.4 and is dependent on concentrations of molybdate and demolybdoenzyme. The reaction is inhibited by high concentrations of anions and by reduction of the demolybdoenzyme and requires incubation temperatures higher than 30 degrees. A reconstitution mechanism involving loss of tungsten and concomitant replacement with molybdenum in those demolybdo molecules which contain tungsten is supported by the following observations: (a) the extent of activation achieved by molybdate corresponds to the proportion of molecules in the preparation which contain tungsten. (b) Incubation of the demolybdoenzyme preparation at 37 degrees in the absence of molybdate results in progressive and concentration-dependent loss of ability to be reconstituted by molybdate and a corresponding but more rapid loss of tungsten from the enzyme. The reconstituted enzyme displays the molybdenum EPR signal characteristic of native enzyme and is inactivated by incubation at 42 degrees in a manner identical to native sulfite oxidase.

Animals↗
PubMed1977

Tryptic cleavage of rat liver sulfite oxidase. Isolation and characterization of molybdenum and heme domains.

Treatment of rat liver sulfite oxidase with trypsin leads to loss of ability to oxidize sulfite in the presence of cytochrome c as electron acceptor. Ability to oxidize sulfite with ferricyanide as acceptor is undiminished, while sulfite leads to O2 activity is partially retained. Gel filtration of the proteolytic products has led to the isolation of two major fragments of dissimilar size derived from sulfite oxidase. The smaller fragment has a molecular weight of 9500 and appears to be monomeric when detached from sulfite oxidase. It contains the heme in its cytochrome b5 structure, has no sulfite oxidase activity, and is reducible with dithionite but not with sulfite. The heme fragment can mediate electron transfer between pig liver microsomal NADH cytochrome b5 reductase and cytochrome c. The larger fragment has a molecular weight of 47,400 under denaturing conditions but elutes from Sephadex G-200 as a dimer. It contains no heme but retains all of the molybdenum and the modified sulfite-oxidizing capacity present in the proteolytic mixture. All of the EPR properties of the molybdenum center of native sulfite oxidase are retained in the molybdenum fragment. The molybdenum center is a weak chromophore with an absorption sectrum suggestive of coordination with sulfur ligands. Reduction by sulfite generates a spectrum attributable to molybdenum (V). Spectra of oxidized and sulfite-reduced preparations are sensitive to anions and pH. NH2-terminal analysis of native sulfite oxidase and the two tryptic fragments has permitted the conclusion that the sequence represented by the heme fragment is the NH2 terminus of native enzyme. These studies have demonstrated that the two cofactor moieties of sulfite oxidase are contained in distinct domains which are covalently held in contiguity by means of an exposed hinge region. Isolation of functional heme and molybdenum domains of sulfite oxidase after tryptic cleavage has demonstrated conclusively that the cytochrome b5 region of the molecule is required for electron transfer to the physiological acceptor, cytochrome c.

Animals↗
PubMed1976

Electron paramagnetic resonance of the tungsten derivative of rat liver sulfite oxidase.

Sulfite oxidase purified from livers of tungsten-treated rats has been used for EPR studies of tungsten substituted at the molybdenum site of the enzyme in a fraction of the molecules. The EPR signal of W(V) in sulfite oxidase is quite similar to that of Mo(V) in its line shape and in its sensitivity to the presence of anions such as phosphate and fluoride. Hyperfine interaction with a dissociable proton is also observed in both signals. The pH-dependent alteration in line shape exhibited by the Mo(V) EPR signal of the rat liver enzyme. Incomplete reduction of the tungsten center at pH 9 is indicated by attenuated signal intensity at this pH. The W(V) signal has g values lower than those of the Mo(V) signal, has a much broader resonance envelope, and is much less readily saturated by increasing microwave power. Kinetic studies on the reduction of the heme and tungsten centers of sulfite oxidase have shown that reduction of de-molybdo forms of sulfite oxidase by sulfite is catalyzed by the residual traces of native molybdenum-containing molecules. Reduction is accomplished by electron transfer involving intermolecular heme-heme interaction. The W(V) signal is generated only after all the heme centers are reduced. The rate and extent of heme reduction at pH 9 are the same as at pH 7. Studies on the reoxidation of W(V) and reduced heme by O2 and by cytochrome c suggest that the cytochrome b5 of sulfite oxidase is the site of electron transfer to cytochrome c, whereas oxidase activity is the property of the molybdenum center. It appears that the tungsten center in sulfite oxidase is incapable of oxidizing sulfite.

Anaerobiosis↗
PubMed1976

Assessment of clinical laboratory techniques in the measurement of oxygen saturation in whole blood.

The accuracy and advantages of determining percentage of hemoglobin oxygen saturation of whole blood by advanced instruments of oximetry, gas chromatography, and the computation of oxygen saturation from pO2 and pH data are compared with the Van Slyke-Neill manometric method. It was found that oximetry is extremely fast, less expensive, and more accurate than in the past, the apparatus requires little training to operate and has diminutive service maintenance. The gas chromatograph proved to be highly accurate and could serve as a standard calibration check method in place of the more time-consuming Van Slyke-Neill apparatus for oximetry and pO2 and pH electrode instrumentation. Computation of O2 saturation from pO2 and pH data also proved to be accurate, confirming that this method is more useful during cardiac surgery because it can provide additional acid-base information. In the low oxygen saturation range (less than 60%), all three methods proved to be of similar sensitivity.

Chromatography, Gas↗
PubMed1976

Silicic acid chromatographic separation of polychlorinated biphenyls and pesticides: some contaminants and limitations.

The Armour and Burke method for separating polychlorinated biphenyls (PCBs) from pesticides involves the use of silicic acid. However, we detected di-2-ethylhexyl phthalate, PCBs, and other impurities as interfering contaminants in several batches of silicic acid. The presence of H2SO4 in some batches of silicic acid is inferred. The acid may be responsible for the production of contaminants which interfere in gas-liquid chromatographic analyses. Contaminants in silicic acid are reduced by extracting the adsorbent with solvent, and/or partitioning the concentrated pesticide fraction with 1N NaOH. These purification procedures provide separations relatively free of impurities, but result in reduced adsorbent activity. PCB-pesticide separations are reproducible only within a given batch lot of silicic acid because of the varying adsorbent characteristics of each lot. Alternative adsorbents should be explored for most PCB-pesticide separations.

Chromatography, Gas↗
PubMed1975

Taxonomy of the Clostridia: ribosomal ribonucleic acid homologies among the species.

rRNA homologies have been determined on reference strains representing 56 species of Clostridium. Competition experiments using tritium-labelled 23S rRNA were employed. The majority of the species had DNA with 27 to 28% guanine plus cytosine (%GC). These fell into rRNA homology groups I and II, which were well defined, and a third group which consisted of species which did not belong in groups I and II. Species whose DNA was 41 to 45% GC comprised a fourth group. Thirty species were placed into rRNA homology group I on the basis of having 50% or greater homology with Clostridium butyricum, C. perfringens, C. carnis, C. sporogenes, C. novyi or C. pasteurianum. Ten subgroups were delineated in homology group I. Species in each subgroup either had high homology with a particular reference species or a similar pattern of homologies to all of the reference organisms. The eleven species in rRNA homology group II had 69% or greater homology to C. lituseburense. Species in groups I and II had intergroup homologies of 20 to 40%. The six species in group II had very low homologies with groups I and II. Negligible homology also resulted when five of the species were tested against the sixth, C. ramosum. The five species having DNA with 41 to 45% GC were C. innocuum, C. sphenoides, C. indolis, C. barkeri and C. orotic um. Little rRNA homology was apparent between C. innocuum and the other high % GC species or with several Bacillus species having similar %GC DNA. Correlations between homology results and phenotypic characteristics are discussed.

Base Sequence↗
PubMed1975

Blood flow velocity in the carotid artery as a measure of myocardial contractility.

An attempt was made to derive a useful noninvasive index to evalute a change in myocardial contractile state using transcutaneous Doppler flow-velocity curve from the carotid artery. In 5 mongrel dogs and in 43 patients with various heart disease, Doppler flow velocity curves were obtained from the ascending aorta intravascularly using a Doppler catheter and/or from carotid artery transcutaneously using a Doppler probe. The first derivative of left ventricular pressure (dp/dt) and electrocardiogram (ECG) were recorded simultaneously. The following 3 indices were measured from the Doppler flow-velocity curves: (1) macimum acceleration of blood flow (dv/dt), (2) time from onset of ejection to peak flow (time-to-peak), (3) time interval between the beginning of Q wave of ECG to the peak of Doppler flow velocity curve (ECG Q-Doppler peak). Among these 3 indices, only ECG Q-Doppler peak demonstrated a significant correlation between the values measured intravascularly and transcutaneously. Also, only ECG Q-Doppler peak showed significant correlation with maximum of dp/dt (max dp/dt). Since ECG Q-Doppler peak showed correlation with heart rate, the difference between observed and predicted ECG Q-Doppler peak (delta ECG Q-Doppler peak) was calculated to exclude the effect of heart rate. Predicted value of ECG Q-Doppler peak was calculated from the regression equation between heart rate and ECG Q-Doppler peak in the separate experiments. There was significant correlation between delta ECG Q-Doppler peak and max dp/dt. In 15 patients with coronary artery disease and in 16 healthy subjects, delta ECG Q-Doppler peak and the other noninvasive method (systolic time intervals) were measured. Delta ECG Q-Doppler peak showed better result in the separation of 2 groups than by systolic timeintervals. It was concluded that delta ECG Q-Doppler peak is a useful index to evaluate the myocardial contractile state since this index is readily obtained noninvasively.

Adolescent↗
PubMedDate not supplied

Bacteriophages of Bacteroides.

Sixty-eight bacteriophages specific for nine species (DNA homology groups) of Bacteroides were isolated from sewage. Four distinct morphological types were isolated, three of which had not previously been described. Attempts to use these phages to transduce Bacteroides fragilis were unsuccessful. A total of 91 phage-susceptible strains of Bacteroides were used with these phages in a study of the feasibility of developing a scheme for identification of Bacteroides species. Blind trials were performed with 10 B. fragilis-specific phages and a collection of over 200 bacterial strains, including 144 strains of B. fragilis. Of the strains of B. fragilis, 78% were identified correctly within 24 hr. A phage-carrier state, or pseudolysogeny, was observed with most of the phage-host systems, and this state was studied in detail with B. fragilis phage Bf-1. The presence of a thick capsule around some cells in a pure culture of a host strain appears to render these cells resistant to phage infection, thus perpetuating the carrier state. It is suggested that such capsules may play a role in the virulence of strains of Bacteroides.

Bacteriophages↗