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Sulfur and methionine metabolism in sheep. I. First approximations of sulfur pools in and sulfur flows from the reticulo-rumen.

Sulfur pools in the rumen and sulfur flows from the rumen were investigated in two experiments with sheep on a diet containing equal parts of oaten and lucerne chaffs. The diet was fed at two levels, either chopped or pelleted, and with intraruminal DL-methionine supplements. Ruminal fluid volumes and fluid flows to the omasum were measured. None of the treatments influenced ruminal fluid volume. Fluid flow to the omasum, however, was increased by increasing dry matter intake (DMI), and was further enhanced by feeding chaffed hay rather than the same materials ground and pelleted; the DL-methionine supplement had no effect. First approximation of the ruminal sulfur pools and of sulfur flows to the omasum were derived from the concentration of sulfur in true digesta and the ruminal fluid volume or fluid flow. Increasing DMI from 500 to 1000 g/day resulted in larger ruminal pools of total (1096 v. 792 mg), neutral (1016 v. 731 mg) and protein (479 v. 419 mg) sulfur, but the reducible sulfur pools were not affected by the level of DMI. Infusions of DL-methionine increased the ruminal sulfide sulfur pool irrespective of level of DMI. The first approximation of total sulfur flow was increased by 1660 mg/day at the higher level of DMI, due mainly to increases of 710 mg S/day as protein sulfur and 859 mg S/day as non-protein neutral sulfur. Flows of inorganic sulfate and ester sulfate sulfur, although small in comparison with organic sulfur flows, increased with level of DMI. Sulfide sulfur flows were also increased at the higher level of DMI, and were almost doubled by intraruminal infusions of DL-methionine.

Animal Feed

MCD spectra of iron-sulfur complexes with or without inorganic sulfur.

The magnetic circular dichroism spectra were observed for various iron-sulfur complexes with and without inorganic sulfur as models for rubredoxin and 2-Fe ferredoxin. The MCD band shapes ascribed the bands around 390 and 490 nm to Faraday A terms for mononuclear iron sulfur complexes. These bands are probably assigned to the charge-transfer transitions from the thiol sulfur orbital to the iron t2 and e 3d-orbitals, respectively. The MCD magnitudes decreased by more than one-half for binuclear iron-sulfur complexes with inorganic sulfur in comparison with those for the mononuclear complexes. The low MCD magnitude as well as the possible core symmetry as low as D2d attributed the MCD bands to Faraday B terms. Incorporation of inorganic sulfur produced new MCD bands, some of which can be assigned to the charge-transfer transitions from the inorganic sulfur orbital to the iron t2 and e 3d-orbitals. Among complexes studied here, the bis(o-xylyldithiolato) ferrate(III) monoanion gave the MCD spectrum which resembles that of a rubredoxin. This implies that the MCD spectroscopy also assessed complex as a good rubredoxin model. However the binuclear complex bis[o-xylyldithiolato-micron2-sulfidoferrate(III)] dianion failed to offer the MCD spectrum similar to that of the spinach ferredoxin.

Circular Dichroism

Sulfur and methionine metabolism in sheep. II. Quantitative estimates of sulfur metabolism in the sheep's stomach.

The metabolism of dietary and supplemental DL-methionine sulfur in the stomach of sheep was studied in two experiments. In both experiments sheep were fed a 50 : 50 oaten chaff: lucerne chaff ration at two levels of intake, and some animals received intraruminal infusions of DL-methionine. In experiment 2 increasing dry matter intake (DMI) increased first approximations of total, neutral, protein and reducible sulfur flows and also sulfide sulfur flow from the reticulo-rumen. Increased DMI (from 500 to 1000 g/day) also resulted in greater true flows of total (2207 v. 1104 mg/day), neutral (1867 v. 1043 mg/day) and protein (893 v. 482 mg/day) sulfur at the duodenum. Two flow diagrams of sulfur metabolism in the compartments of the ruminant stomach were developed from the data of experiment 2. Fluid flows of sulfur in experiment 1 were used to supplement the data of experiment 2 in developing the balance models. The two models represent the extremes of dietary and supplemental sulfur metabolism in the sheep's stomach under the conditions of experiment 2, and they are discussed in relation to previous research on sulfur metabolism in the stomach.

Animals

Effect of sulfur application on sulfur-oxidizing bacteria and yield of two leguminous crops.

The effect of elemental sulfur application at the rate of 50, 100, 200, and 400 kg/feddan on broad-bean and lentil was tested in field experiments. Sulfur application resulted in increasing the numbers of the autotrophic sulfur-oxidizing bacteria as well as in the concentration of available sulfate in the soil; the increases were proportional to the level of sulfur added. The sulfur treatments neither affected the soil pH nor the nodulation of the leguminous plants tested. The application of sulfur at the rates used slightly increased seed yield and nitrogen content of lentil seeds, but had no effect on broadbean.

Fabaceae

Long-term exposure to sulfur dioxide, sulfuric acid mist, fly ash, and their mixtures. Results of Studies in Monkeys and guinea pigs.

Groups of cynomolgus monkeys and guinea pigs were exposed to mixtures of sulfur dioxide, fly ash, and sulfuric acid mist. The exposure concentrations varied between 0.1 and 5.0 ppm for sulfur dioxide, 0.1 and 1 mg/cu m for sulfuric acid mist, while a concentration of approximately 0.5 mg/cu m was used for fly ash. The duration of exposure was 52 weeks for guinea-pigs and 78 weeks for monkeys. Pulmonary function tests and serum biochemical and hematological analyses were conducted prior to and periodically during the exposure period. At the termination of exposure, the lungs were examined microscopically. Analysis of the data revealed that in groups exposed to the mixtures of pollutants, sulfuric acid mist was responsible for the effects observed. No synergistic action between the pollutants was detected.

Aerosols

The high potential iron-sulfur cluster of aconitase is a binuclear iron-sulfur cluster.

It has been reported (Ruzicka, F.J., and Beinert, H. (1978) J. Biol. Chem. 253, 2514-2517) that aconitase in the oxidized state, as isolated, shows an electron paramagnetic resonance signal centered at g = 2.01, typical of high potential iron-sulfur proteins. Since the magnetic state corresponding to this signal has thus far only been found in tetranuclear iron-sulfur clusters in model compounds and proteins, it could be expected that aconitase also contains a [4Fe-4S] cluster. We show here that core extrusion, in the presence of hexamethylphosphoramide and o-xylyl-alpha,alpha'-dithiol and subsequent ligand exchange with p-trifluoromethylbenzenethiol yield absorption spectra typical of binuclear iron-sulfur clusters. According to the absorbance measured, the concentration of the extruded [2Fe-2S] cluster quantitatively accounts for the iron-sulfur content of the preparations examined. Preliminary studies of the 19F nuclear magnetic resonance spectrum obtained on extrusion with p-trifluoromethylbenzenethiol confirm the presence of a binuclear cluster in aconitase.

Aconitate Hydratase

[Comparative study of B890 pigment-lipoprotein complexes from sulfur (Chromatium minutissimum) and non-sulfur (Rhodopseudomonas palustris) purple photosynthesizing bacteria].

Pigment-lipoprotein B890 complexes containing reaction center and "light-focusing" bacteriochlorophyll a were isolated from photosynthetic membranes of sulfur (Chromatium minutissimum) and non-sulfur (Rhodopseudomonas palustris) purple bacteria after the treatment with Triton X-100. The molecular weights of complexes were evaluated using several methods (200 000-300 000). By means of electron microscopy the sizes of complexes were found to be about 80 A. On the air-water interface hexagonal packing of complexes was observed. The chemical compositions of complexes are very similar except bacteriochlorophyll a whose specific content is somewhat higher in Chromatium minutissimum. The protein composition of complexes was studied and the molecular weights of proteins were estimated by SDS-gel electrophoresis. The results obtained show significant similarities in molecular organization of B890 complexes isolated from sulfur (Chromatium minutissimum) and non-sulfur (Rhodopseudomonas palustris) purple bacteria.

Bacterial Proteins

The use of stable sulfur isotope labelling to elucidate sulfur metabolism by Clostridium pasteurianum.

An unique isotope labelling experiment was conducted whereby mixtures of sulfate and sulfite of different isotopic compositions were metabolized by Clostridium pasteurianum. The results showed during reduction of 1 mM SO3= plus 1 mM SO4=, essentially all evolved H2S arose from the sulfite whereas in the case of cellular sulfur, 85% was derived from sulfite and the remainder from sulfate.

Clostridium

Stable isotope fractionation by Clostridium pasteurianum. 2. Regulation of sulfite reductases by sulfur amino acids and their influence on sulfur isotope fractionation during SO32- and SO42- reduction.

In addition to an assimilatory sulfite reductase, studies of cultures of Clostridium pasteurianum supplemented with methionine, cysteine, and 35SO42- provides evidence for another reductase which is induced by SO32-. This inducible reductase appears to be dissimaltory because of the copious sulfide production arising when the cells are grown on SO32-. Cysteine can repress the assimilatory sulfite reductase but does not affect the inducible reductase. During late logarithmic growth on 1 mM SO42- + 10mM cysteine, depression of the inducible reductase occurred along with increased sulfide production. The presence of 1 mM cysteine and (or) 1 mM cysteine and (or) 1 mM methionine does not affect the inverse sulfur isotope effect for evolved H2S. However, 5 and 10 mM cysteine reduce the maximum delta34S value for released H2S from +40 to 10%. A small conversion of cysteine to H2S by C. pasteurianum occurs, but only in the stationary phase.

Cell-Free System

Isolation of the domain containing the molybdenum, iron-sulfur I, and iron-sulfur II centers of chicken liver xanthine dehydrogenase.

Chicken liver xanthine dehydrogenase, like other xanthine-oxidizing enzymes, is a dimer of Mr = 150,000 subunits. Each subunit contains one molybdenum, one FAD, and two distinct Fe2S2 centers. Treatment with a number of proteases shows that the native enzyme subunit is cleaved at three distinct sites. However, the cleavage products can be separated only under denaturing conditions. Prolonged treatment with subtilisin at pH 10.1 has permitted the isolation of an Mr = 65,000 catalytically active fragment that is devoid of FAD but which contains the molybdenum and both types of iron-sulfur center. A model of the domain structure of the native enzyme is proposed.

Animals

[Fractionation of sulfur isotopes by phototrophic sulfur bacterium Ectothiorhodospira shaposhnikovii].

Two processes of sulphur isotope fractionation have been found in experiments with the sulphur purple bacterium Ectothiorhodospira shaposhnikovii. As a result, a light isotope, 32S, is concentrated in residual hydrogen sulphide, and a heavy isotope, 34S, in elementary suphur which is deposited outside the cell. The sulphate produced is lighter than elementary sulphur. Fractionation of sulphur isotopes is observed in natural conditions and is confined to places of mass growth of photosynthetic sulphur bacteria.

Chemical Fractionation