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G Fuchs

Publications and source records attributed to G Fuchs.

At least 37 records · Page 2Linked to original sources

Lipoprotein-induced modulation of cyclosporine-A-mediated immunosuppression.

Human serum lipoproteins form complexes with cyclosporine-A and act as a carrier of cyclosporine-A in vivo. We compared the immunosuppressive effects of free cyclosporine-A, a complex composed of cyclosporine-A and lipoproteins, free cyclosporine-A in the presence of each unbound lipoprotein, and each lipoprotein without cyclosporine-A with one another at concentrations comparable with in vivo conditions on PHA-stimulated peripheral blood mononuclear cells. Free cyclosporine-A reduced the proliferation of the PHA-stimulated mononuclear cells to 50% at a concentration of 300 ng ml-1 (SD +/- 30, n = 12) lipoprotein-deficient medium. Cyclosporine-A loaded into VLDL showed a 50% proliferation rate reduction at 60 micrograms VLDL ml-1 (SD +/- 10, n = 12) and 180 ng cyclosporine-A ml-1. In the presence of 100 ng ml-1 cyclosporine-A 180 micrograms ml-1 VLDL (SD +/- 25, n = 12) showed a proliferation rate reduction of 50%. In the same way VLDL without cyclosporine-A induced a reduction to 50% at 740 micrograms ml-1 (SD +/- 30, n = 12). Cyclosporine-A loaded into LDL showed a 50% proliferation rate reduction at 27 micrograms ml-1 LDL (SD +/- 5, n = 12) with 80 ng ml-1 cyclosporine-A. In the presence of 100 ng ml-1, cyclosporine-A 150 micrograms ml-1 LDL (SD +/- 25, n = 12) showed a proliferation rate reduction of 50%. In the same way, LDL without cyclosporine-A induced a reduction to 50% at 950 micrograms ml-1 (SD +/- 50, n = 12).(ABSTRACT TRUNCATED AT 250 WORDS)

Cyclosporins

13C-NMR study of acetate assimilation in Thermoproteus neutrophilus.

Acetate assimilation into amino acids and the functioning of central biosynthetic pathways in the extremely thermophilic anaerobic archaebacterium Thermoproteus neutrophilus was investigated using 13C NMR as the method for determination of the labelling patterns. Acetate was assimilated via reductive carboxylation of acetyl-CoA to pyruvate and pyruvate conversion to phosphoenolpyruvate which was further carboxylated to oxaloacetate. 2-Oxoglutarate was mainly formed via citrate. However, the labelling patterns of glutamic acid and alanine were in agreement with the concurrent synthesis of about 15% 2-oxoglutarate and 5% pyruvate through the reductive citric acid cycle. A scrambling phenomenon occurring in aspartate and all amino acids derived through oxaloacetate was observed. The labelling patterns of amino acids were in agreement with their standard biosynthetic pathways, with two remarkable exceptions: isoleucine was synthesized via the citramalate pathway and lysine was synthesized via the 2-aminoadipate pathway which has previously been reported only in eukaryotic microorganisms.

Acetates

2-Aminobenzoyl-CoA monooxygenase/reductase, a novel type of flavoenzyme. Purification and some properties of the enzyme.

A novel aerobic mechanism of 2-aminobenzoate metabolism was proposed in a denitrifying Pseudomonas species. 2-Aminobenzoic acid is activated in a coenzyme-A-ligase reaction to 2-aminobenzoyl-CoA and this intermediate is dearomatized by a unique enzyme, tentatively named 2-aminobenzoyl-CoA monooxygenase/reductase. This paper describes the purification and some molecular, kinetic and spectral properties of this flavoenzyme which catalyzes the hydroxylation and reduction of 2-aminobenzoyl-CoA to an unknown non-aromatic compound. 2-Aminobenzoyl-CoA monooxygenase/reductase was purified 25-fold to a specific activity of 25 mumol.min-1.mg-1 protein using ammonium sulfate precipitation, DEAE-cellulose anion-exchange, hydroxylapatite and Mono Q FPLC anion-exchange chromatography. Superose 6 gel filtration for estimation of molecular mass resulted in one symmetrical protein peak corresponding to a molecular mass of 170 kDa. Several experimental data suggest that the protein is probably an alpha 2 dimer; however, it may exist in three dimeric forms, alpha alpha, alpha alpha' and alpha' alpha', where alpha' may be a subunit with a different conformation. Approximately 2 mol noncovalently bound FAD/mol enzyme was found, which in the absence of O2 was reduced by NADH. The enzyme was specific for the substrates 2-aminobenzoyl-CoA (Km less than or equal to 25 microM) and O2 (Km less than or equal to 5 microM), but less specific for the reduced pyridine nucleotides NADH (Km = 42 microM) or NADPH [Km = 500 microM; Vmax (NADH)/Vmax (NADPH) = 1.7:1]. The turnover number was 4250 min-1. The enzyme also reduced N-ethylmaleimide and maleimide with NAD(P)H. The substrate, the products and the reaction stoichiometry are described in two following papers.

Catalysis

2-Aminobenzoyl-CoA monooxygenase/reductase, a novel type of flavoenzyme. Studies on the stoichiometry and the course of the reaction.

The reaction catalyzed by 2-aminobenzoyl-coenzyme-A monooxygenase/reductase from a denitrifying Pseudomonas sp. has been investigated. 2-Aminobenzoyl-CoA and 2-amino[carboxy-14C]benzoyl-CoA were synthesized enzymatically using 2-aminobenzoyl-CoA synthetase from the same organism. The product was purified by chromatography and characterized by ultraviolet/visible and 1H-NMR spectroscopy. The conversion of 2-aminobenzoyl-CoA catalyzed by the monooxygenase/reductase requires NADH and oxygen, and yields at least two different products depending on the relative concentration of NADH. At [NADH] less than Km (40 microM), i.e. [NADH]/[2-aminobenzoyl-CoA] approximately 0.02-0.05, the main product is probably a hydroxylated derivative of 2-aminobenzoyl-CoA, which is characterized by an absorbance maximum around 375 nm. When [NADH]/[2-aminobenzoyl-CoA] approximately 2-5, the predominant product is a non-aromatic coenzyme A thioester (lambda max approximately 320 nm). The stoichiometry in this case is 2.1-2.4 mol NADH oxidized (mol oxygen consumed)-1 (mol 2-aminobenzoyl-CoA metabolized)-1. The product is extremely unstable in the acidic pH range and undergoes decarboxylation in a few minutes at pH less than 5. Some degree of stabilisation is obtained upon reduction with sodium borohydride, probably resulting in a further reduced non-aromatic coenzyme-A thioester.

Catalysis

13C-NMR study of autotrophic CO2 fixation in Thermoproteus neutrophilus.

The pathway of autotrophic CO2 fixation has been investigated in the extremely thermophilic sulfur-respiring anaerobic archaebacterium Thermoproteus neutrophilus. [1,4-13C2]Succinate was used as a tracer since this compound was incorporated in small amounts virtually into all cell compounds without affecting the organism's ability to synthesize all cell constituents from CO2. Three representative amino acids, glutamate, aspartate and alanine were isolated from cells after growth for several generations in the presence of [1,4-13C2]succinate and their labelling patterns were determined by 13C NMR spectroscopy. The data is consistent with CO2 fixation by a reductive citric acid cycle, as proposed earlier for the green sulfur bacterium Chlorobium limicola, the sulfate-reducing Desulfobacter hydrogenophilus and the microaerophilic Knallgasbacterium Hydrogenobacter thermophilus. The presence of a reductive citric acid cycle in archaebacteria indicates that this CO2 fixation mechanism which is an alternative to the Calvin cycle is present in many anaerobic or facultative anaerobic microorganisms.

Archaea

Reductive dehydroxylation of 4-hydroxybenzoyl-CoA to benzoyl-CoA in a denitrifying, phenol-degrading Pseudomonas species.

The initial reactions in anaerobic degradation of phenol to CO2 have been studied in vitro with a denitrifying Pseudomonas strain grown with phenol and nitrate in the absence of molecular oxygen. Phenol has been proposed to be carboxylated to 4-hydroxybenzoate [(1987) Arch. Microbiol. 148, 213-217]. 4-Hydroxybenzoate was activated to 4-hydroxybenzoyl-CoA by a coenzyme A ligase. Cell extracts also catalyzed the reductive dehydroxylation of 4-hydroxybenzoyl-CoA to benzoyl-CoA with reduced benzyl viologen as electron donor. This enzyme, benzoyl-CoA:(acceptor) 4-oxidoreductase (hydroxylating) (EC 1.3.99.-), has not been reported before. The data suggest that phenol and 4-hydroxybenzoate are anaerobically metabolized by this strain via benzoyl-CoA.

Acyl Coenzyme A

Diurnal variations of blood pressure in shift workers during day and night shifts.

The dependence of blood pressure upon internal rhythms and the short-term effects of shift rota on the blood pressure were investigated in shift workers. Blood pressure was measured every 30 min using automatic records for 24h in 17 physically working men in a chemical factory during their morning and night shifts. There were no differences of the mean blood pressure between the respective sleeping phases or between the working periods. The amplitudes of circadian blood pressure variations were equal. There was a phase difference of 8h corresponding to the lag between the working periods. At this 8-h lag the hourly means of the 24-h blood pressure were closely correlated (r=0.69). Comparisons of 24-h blood pressure profiles during the first and last days of night shift week showed that the effects of night work on the blood pressure were already fully developed within the first 24h (r=0.86). Thus the diurnal variations of the blood pressure are determined by the working and sleeping periods and largely independent of endogenous rhythm. There is no short-term alteration of the mean 24-h blood pressure after shift rota.

Adult

Enzyme reactions involved in anaerobic cyclohexanol metabolism by a denitrifying Pseudomonas species.

The enzymes involved in the anaerobic degradation of cyclohexanol were searched for in a denitrifying Pseudomonas species which metabolizes this alicyclic compound to CO2 anaerobically. All postulated enzyme activities were demonstrated in vitro with sufficient specific activities. Cyclohexanol dehydrogenase catalyzes the oxidation of the substrate to cyclohexanone. Cyclohexanone dehydrogenase oxidizes cyclohexanone to 2-cyclohexenone. 2-Cyclohexenone hydratase and 3-hydroxycyclohexanone dehydrogenase convert 2-cyclohexenone via 3-hydroxycyclohexanone into 1,3-cyclohexanedione. Finally, the dione is cleaved by 1,3-cyclohexanedione hydrolase into 5-oxocaproic acid. Some kinetic and regulatory properties of these enzymes were studied.

Aerobiosis

Nephrolithiasis.

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Humans

[Extracorporeal shock wave lithotripsy: the evolution of a revolution].

Extracorporeal shockwave lithotripsy has now been in clinical use for 8 years, and it has replaced other treatment techniques for the majority of surgical calculi in the upper urinary tract. For the first time it provides a completely noninvasive method for the treatment of renal and ureteral calculi. The current range of indications means that approximately 70% of nonselected urinary stone patients can be treated by this method, while 25% of the patients with more complex stones in the upper urinary tract can receive treatment with the lithotripter combined with endourological procedures. The clinical role of this method is determined by a high success rate and minimal complications. This has led to a rapid worldwide acceptance in the urological community, and up to now more than 1.5 million patients with urinary stone disease have been treated.

Combined Modality Therapy

Twenty-four-hour blood pressure is not dependent on endogenous circadian rhythm.

The effects of shifted working and sleeping phases on the diurnal blood pressure rhythm were investigated in 15 physically working industrial shift workers at a slowly rotated three-shift system. Ambulatory 24-h blood pressure monitoring was performed during the morning and night shifts. In the two shifts the mean 24-h blood pressure was identical. There were no differences in the blood pressure levels in the sleeping phases or in the working periods between the two 24-h cycles. Diurnal blood pressure fluctuations had equal amplitudes. Corresponding to the lag between the working period there was a phase difference of 8 h between the 24-h blood pressure curves. At this lag, there was a high correlation between the mean hourly blood pressure values (r = 0.683). Twenty-four-hour blood pressure curves during the first and last day of a night shift were nearly equal. Thus the effects of shift rotation on the 24-h blood pressure profile were fully expressed within the first 24 hours. The immediate and complete adaptation of the 24-h blood pressure curve to shifted activity and sleeping phases indicates that activity determines the diurnal blood pressure profile. The blood pressure is largely independent of internal circadian rhythm.

Adaptation, Physiological

Purification and some properties of the corrinoid-containing membrane protein from Methanobacterium thermoautotrophicum.

The cytoplasmic membrane of the methanogenic archaebacterium Methanobacterium thermoautotrophicum does not contain cytochromes, but did contain a corrinoid protein of molecular mass about 33 kDa which, after treatment with 10 mg Triton X-100/mg protein, was contained in a protein complex of about 500 kDa. Washed membranes from 1 g dry cells contained about 70 nmol of the cobamide factor III (5-hydroxybenzimidazolyl cobamide) as the sole corrinoid. The corrinoid-containing protein complex was purified and some of its properties were studied. According to several criteria it is an integral membrane protein complex. The corrinoid-protein complex, after about 100-fold purification, gave a single band on native PAGE and still had molecular mass of about 500 kDa. In SDS-PAGE several subunits were observed: in addition to the corrinoid-carrying subunit of about 33 kDa, other polypeptides of approximately 28 kDa, 26 kDa, and possibly 23 kDa were present. One mole of the purified 500-kDa protein complex contained greater than or equal to eight moles of the cobamide factor III. It was estimated that the corrinoid-protein complex accounts for 8% of the membrane protein of M. thermoautotrophicum. The visible spectrum of the oxidized protein exhibited absorbance maxima at 547 nm, 511 nm, and a shoulder at 468 nm, which disappeared upon reduction with dithionite. The midpoint potential of this transition was around -145 mV (pH 7). With EPR a Co2+ signal was observed within -50 mV and -350 mV with a maximum around -200 mV. Possible reasons for the disappearance of the Co2+ signal at low redox potentials are discussed. The line shape of the Co2+ signal was similar to that of Co2+ in free corrinoids. The signal of Co2+ could also be evoked by reduction with 5 mM dithiothreitol. From the redox properties of the corrinoid membrane protein it may be expected that in vivo the cobalt may become reduced and reoxidized. Its possible function as an electron-mediating membrane protein in the metabolism of methanogenic bacteria is discussed.

Bacterial Proteins

Anaerobic metabolism of cyclohexanol by denitrifying bacteria.

Three strains of denitrifying bacteria were anaerobically enriched and isolated from oxic or anoxic habitats with cyclohexanol or cyclohexanone as sole electron donor and carbon source and with nitrate as electron acceptor. The bacteria were facultatively anaerobic, Gram-negative and metabolism was strictly oxidative with molecular oxygen, nitrate, or nitrite as terminal electron acceptor. Cyclohexanol and cyclohexanone were degraded both anaerobically and aerobically. Aromatic compounds were oxidized in the presence of molecular oxygen only. One of the bacterial strains was further characterized. During anaerobic cyclohexanol degradation approximately 40% of the substrate was oxidized to phenol, which accumulated as dead-endproduct in the growth medium; 60% of cyclohexanol was completely oxidized to CO2 and assimilated, respectively. In addition to phenol formation, transient accumulation of cyclohexanone, 2-cyclohexenone and 1,3-cyclohexanedione was observed. Based on these findings we propose a pathway for anaerobic cyclohexanol degradation involving these intermediates.

Anaerobiosis

Open comparative study with treatment-refractory depressed patients: electroconvulsive therapy--anesthetic therapy with isoflurane (preliminary report).

Isoflurane is the only routine volatile anesthetic which leads to a burst-suppression EEG in nontoxic concentrations and for which no memory impairments can be expected subsequent to anesthesia. After electrically induced spike-wave activity in electroconvulsive therapy (ECT), a brief period of a very flat EEG can be observed. This phenomenon may be closely related to the therapeutic effects of ECT. The results which have been achieved up to now in comparing ECT to anesthetic therapy with isoflurane, without seizures being induced, are presented.

Adult