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G Cecchini

Publications and source records attributed to G Cecchini.

At least 73 records · Page 4Linked to original sources
PubMed1986

Age related changes of glucagon binding and activity in isolated rat hepatocytes.

Hepatocytes isolated from young and old rats were compared for glucagon binding and its biological effect. Liver cells from young rats showed a receptor binding of the hormone significantly higher than cells from old rats. Such a behaviour was paralleled by the activity of the hormone. In fact, the glucagon-dependent stimulation of adenylate cyclase in the hepatocytes from the former group of animals was twice as high as in the latter.

Adenylyl Cyclases↗
PubMed1985

The high potential iron-sulfur center in Escherichia coli fumarate reductase is a three-iron cluster.

The fumarate reductase complex and soluble enzyme from Escherichia coli have been investigated by low temperature magnetic circular dichroism and electron paramagnetic resonance spectroscopies. The results confirm the presence of one [2Fe-2S] cluster and show that the high potential iron-sulfur center is a 3Fe cluster of the type found in bacterial ferredoxins. Since the 3Fe cluster is present in catalytically competent enzyme and does not appear to be involved in any type of cluster conversion under reducing conditions, we conclude that it is an intrinsic component of the functional enzyme. The significance of the results is discussed in relation to the published amino acid sequence and the iron-sulfur cluster composition of bacterial fumarate reductases.

Amino Acid Sequence↗
PubMed1985

In vivo detection of a three iron cluster in fumarate reductase from Escherichia coli.

Escherichia coli with plasmid amplified expression of fumarate reductase was grown anaerobically on a medium containing fumarate and glycerol and investigated by electron paramagnetic resonance spectroscopy. Anaerobically harvested cells exhibit an EPR signal characteristic of a reduced [2Fe-2S] cluster. Anaerobic addition of fumarate results in diminution of the reduced [2Fe-2S] signal and the appearance of the EPR signal associated with the oxidized 3Fe cluster. The results provide the first evidence for a trinuclear iron-sulfur cluster that exists in vivo, and suggest that the 3Fe cluster in purified fumarate reductase samples is not an artifact of the isolation procedure. The significance of this observation is discussed in relation to the physiological relevance of trinuclear iron-sulfur clusters.

Circular Dichroism↗
PubMed1985

Detection of a tetranuclear iron-sulfur center in fumarate reductase from Escherichia coli by electron paramagnetic resonance spectroscopy.

Soluble fumarate reductase and fumarate reductase complex from Escherichia coli have been investigated by electron paramagnetic resonance spectroscopy. Both succinate- and dithionite-reduced samples show signals associated with a [2Fe-2S]1+ cluster that account maximally for slightly more than one spin/molecule. In addition, at temperatures below 20 K, dithionite-reduced samples exhibit broad and complex features, to high and low field of the [2Fe-2S]1+ signal, that are attributable to a spin coupled [4Fe-4S]1+ cluster. Preliminary attempts to quantify the signals indicate that the [4Fe-4S] cluster is present in an approximate 1:1 stoichiometry with the [2Fe-2S] cluster. The observed enhancement of the spin relaxation of the [2Fe-2S]1+ cluster on dithionite reduction is attributed to spin-spin interaction between the S = 1/2, reduced tetranuclear and binuclear clusters.

Dithionite↗
PubMed1985

The biosynthesis and assembly of methanol dehydrogenase in bacterium W3A1.

Bacterium W3A1, a restricted facultative methylotroph, produces a periplasmic methanol dehydrogenase composed of two identical subunits of Mr = 57,300, and two noncovalently bound methoxatin prosthetic groups. A precursor form of Mr = 1,500 larger than the mature subunit was identified among the products of an in vitro translation of total RNA isolated from bacterium W3A1. The precursor form of the protein could not be detected in cells during in vivo pulse-labeling studies, suggesting that the processing of this precursor occurs entirely co-translationally. Whereas the holoenzyme was detectable only as a dimer, removal of the prosthetic group yielded an apoenzyme that could be detected as either a dimeric or monomeric species. After readdition of the purified prosthetic group to the apoenzyme, only the dimeric form of the protein, bearing the cofactor and exhibiting an absorption spectrum similar to that of the holoenzyme, was detected. Neither the mature apoprotein nor the holoenzyme demonstrated any affinity for phospholipid membranes, as assayed by their inability to bind to liposomes. Taken together, these data suggest a scheme of co-translational processing and export of the apoprotein subunits, followed by assembly of the subunits and prosthetic groups in the periplasmic space to form the mature holoenzyme. The suitability of bacterium W3A1, and other methylotrophic bacteria, for use in studies of protein biosynthesis and export, is also discussed.

Alcohol Oxidoreductases↗
PubMed1984

Further experiments on lipid peroxidation in transplanted and experimental hepatomas.

The results of experiments on the subject of lipid peroxidation in hepatomas are described. It is now clear that lipid peroxidation is strongly decreased in most highly dedifferentiated hepatomas. It seems evident that the extent of the decline is strictly related to the degree of dedifferentiation. The model of diethylnitrosamine carcinogenesis, according to the method by Solt, Medline and Farber, has been now adopted to study the stages of carcinogenesis. It was shown that a net decline in lipid peroxidation occurs as early as at the stage of reversible nodules and progresses until the development of clear hepatomas. This change is practically simultaneous with a decline in the efficiency of the enzymes of the drug metabolizing system and in the content of cytochrome P450-Glutathione content and metabolism show also important changes. In fact, a dramatic increase in gamma-glutamyl-transpeptidase takes place very early during carcinogenesis, and is responsible for large decline in total glutathione during incubation of the homogenates. Glutathione peroxidase activity, on the contrary, is decreased, whereas glutathione reductase does not show significant changes. The supernatant of highly anaplastic tumors inhibits lipid peroxidation in normal liver homogenates, suggesting the presence of substances provided with antioxidant properties. These cannot be, however, related to a higher glutathione content. Supernatants from early nodules seem to be unable to block lipid peroxidation in normal liver homogenates. Preliminary experiments done to study the aldehyde pattern produced during lipid peroxidation, both in hepatomas and in nodules, confirm the presence of very poor lipid peroxidation and possibly of different peroxidation kinetics.

Aminopyrine N-Demethylase↗
PubMed1983

Lipid peroxidation in hepatomas of different degrees of deviation.

Lipid peroxidation rate in four different hepatomas is quite different and seems to be related to their degree of deviation, low deviation tumours displaying higher peroxidative ability. Moreover, the supernatant of the highly anaplastic Yoshida hepatoma is able to decrease the peroxidation rate in normal liver microsomes. This antioxidant ability is not dependent upon an increased level of glutathione. The concentration of reduced glutathione (GSH) declines strongly during incubation in conditions favouring lipid peroxidation. Unlike normal liver homogenates, this decline of GSH in hepatomas is not due to the transformation of GSH into oxidized glutathione (GSSG) but mostly to the increased activity of the gamma-glutamyl-transpeptidase pathway.

Animals↗
PubMed1983

Biochemical evidence for chemical and/or topographic differences in the lipoperoxidative processes induced by CCl4 and iron.

Isolated rat hepatocytes, treated with CCl4 or ADP-Fe3+ complex show an enhanced lipid peroxidation and a decreased glucose 6-phosphatase activity. Lipid peroxidation is much more stimulated by ADP-Fe3+ or Fe3+ than by CCl4, when the metal and the haloalkane are used at a similar concentration. Increasing rates of lipid peroxidation in the different experimental conditions do not correlate with the degree of glucose 6-phosphatase inactivation, which is produced by CCl4 and not by a similar amount of ferric iron. In the case of iron, its intracellular concentration must be higher to give the enzyme inactivation exerted by CCl4. Higher intracellular levels of iron are reached when the metal is added to the cell suspension together with ADP. Under these conditions there is inactivation of glucose 6-phosphatase. Possible mechanisms accounting for a different enzyme sensitivity to iron and CCl4 are discussed.

Adenosine Diphosphate↗
PubMed1982

Functional impairment of intact rat liver cells due to biological aldehydes.

The addition of lipid peroxidation end-products, 4-hydroxynonenal (HNE) or hexanal (HEX) to the incubation medium of rat hepatocytes caused significant decrease of cell cytochrome P-450 content and inactivation of total cell glucose-6-phosphatase. Both the tested aldehydes exerted a marked inhibition of triglyceride secretion by liver cells. The reported results on intact cells furtherly support a possible damaging effect of aldehydes in pathological conditions in which a stimulation of lipid peroxidation occurs.

Aldehydes↗
PubMed1982

HPLC analyses of the aldehydic patterns produced during lipid peroxidation.

The peroxidative breakdown of membrane lipids leads to the production of several compounds with carbonyl functions. Their production has been monitored in rat liver homogenates treated with prooxidant agents namely vitamin C and ADP-Fe2+ complex. Preliminar HPLC separation of the unpolar and medium polar carbonyls produced, allowed the identification of few of them, in particular, hexanal and 4-OH-nonenal. The aim of this report is to underline the usefulness to integrate malonaldehyde evaluation with the analysis of the other carbonyls produced during lipid peroxidation.

Adenosine Diphosphate↗
PubMed1982

Control of cell protein catabolism in rat liver. Effects of starvation and administration of cycloheximide.

1. The loss of liver protein occurring in rats starved for 24 h was largely prevented by the administration of repeated doses of cycloheximide, an inhibitor of protein synthesis. Similar effects were produced on tubulin, a 'fixed' liver protein. 2. Starvation accelerated, whereas cycloheximide markedly lowered, the rate of protein radioactivity decay after labelling with [3H]valine or [14C]bicarbonate, indicating that changes in catabolic rates played an important role in the above regulations of liver protein mass. 3. The total activity of several lysosomal hydrolases showed little change in livers of starved rats, but a marked progressive decline developed after the administration of cycloheximide, particularly in the activities of cathepsins B, D and L as well as acid ribonuclease. There was no evidence that these changes might be due to endogenous inhibitors (at least for cathepsin B activity, which fell to less than 30% of the control values) or enzyme leakage into the bloodstream; rather, plasma beta-galactosidase and beta-N-acetylglucosaminidase activities fell progressively during the cycloheximide treatment. 4. Endogenous proteolytic rates, measured in vitro by incubating subcellular preparations from livers prelabelled in vivo with [3H]valine, were markedly decreased in cycloheximide-treated animals. 5. The osmotic fragility of hepatic lysosomes, appreciably enhanced in starved animals, after cycloheximide treatment was found to be even lower than in fed controls. 6. The present data are consistent with the view that in starved animals the loss of liver protein is mostly accounted for by increased breakdown, due, in part at least, to enhanced autophagocytosis. 7. Cycloheximide largely counteracted these effects of starvation, altering the liver from being 'poised' in a proteolytic direction to a protein-sparing condition. The present data suggest that, besides suppression of the autophagic processes, a decrease in the lysosomal proteolytic enzyme system may also play a role in this regulation, and they seem to provide further circumstantial evidence for the existence of co-ordinating mechanisms between protein synthesis and degradation.

Animals↗
PubMed1981

[Differences in glutathione oxidation and transpeptidylation between normal liver and hepatomas (author's transl)].

Total homogenates from liver tissues, as well from Morris 3924 A and Yoshida AH-I30 hepatomas display a different degree of thiobarbituric acid reacting substances (TBArs) when incubated "in vitro". It is well known that carbonyl compounds arising from lipoperoxidative decomposition of unsaturated fatty acids can easily react with reduced glutathione (GSH). So, the decay in GSH we have shown in previous experiments could be accounted for GSH trapping by the formed aldehydes. Some discrepancies were, however, seen when the decay in GSH and the increase in GSSG were compared, both in normal and in tumour tissues. It is known that GSH can be destroyed not only through oxidative process, but also through the action of gamma-glutamyl-transpeptidase. In the present paper the decrease of total (TG) and reduced (GSH) glutathione was followed and compared with both the increase in GSSG and the increase in the production of TBArs, during "in vitro" incubation. In normal liver, increase in TBArs production parallels the decay in GSH concentration; GSSG, on the contrary, increases. In AH-I30 Yoshida hepatoma cells, TBArs production is lower and GSSG is also decreased. In 3924 A Morris hepatoma GSH decrease is similar to that observed in the liver, while TBArs production is lower and GSSG is also decreased. Analysis of TG content during the incubation-time suggests that GSH decay in both hepatoma types is essentially due to gamma-glutamyl-transpeptidase action, whilst GSH oxidation to GSSG is decreased.

Animals↗
PubMed1980

Uptake and binding of riboflavin by membrane vesicles of Bacillus subtilis.

Riboflavin uptake and membrane-associated riboflavin-binding activity have been investigated in Bacillus subtilis. The uptake and binding activity of the vitamin were found to be repressed coordinately by riboflavin present in the growth medium. The uptake or riboflavin has been shown to have properties of a carrier-mediated process, and membrane vesicles have been shown to demonstrate riboflavin counterflow and exchange. The membrane-associated binding activity for riboflavin has been solubilized with detergents, and a procedure for the partial purification of this component is described. The partially purified riboflavin-binding component has properties expected for a carrier involved in riboflavin uptake, as it shows saturation kinetics and is inhibited by riboflavin analogues. Evidence is also presented showing that reduced riboflavin binds to a greater extent than oxidized riboflavin, and the possible role of the reduced riboflavin in riboflavin uptake is discussed.

Bacillus subtilis↗
PubMed1979

Transport and binding of riboflavin by Bacillus subtilis.

Riboflavine uptake and membrane-associated riboflavin-binding activity has been investigated in Bacillus subtilis. Riboflavin uptake proceeds via a system whose general properties are indicative of a carrier-mediated process: it is inhibited by substrate analogues, exhibits saturation kinetics, and is temperature-dependent. The organism concentrates riboflavin primarily as the phosphorylated cofactors FMN and FAD. Energy is required for uptake but whether the energy demand is required for both uptake and phosphorylation or only for the phosphorylation step is not known. Membrane-associated binding activity for riboflavin has also been demonstrated in membrane vesicles prepared from B. subtilis, and the binding component can be "solubilized" with Triton X-100. Evidence supporting the function of the binding component in riboflavin uptake by the intact cells includes the following. (i) Riboflavin analogues inhibit binding and uptake to nearly the same extent and with similar specificity of action. (ii) The KD for riboflavin-binding and the Km for uptake are in the same range. Similarly the Ki determined for the inhibitory analogue 5-deazariboflavin in the uptake assay and the KD for its interaction with the riboflavin-binding component of membrane vesicles are in the same range. (iii) Uptake in cells and binding in vesicles vary in the same direction with differences in growth conditions.

Bacillus subtilis↗
PubMed1979

Regulation of endogenous proteolysis in rat liver slices.

Methodological difficulties limit studies on cell protein catabolism both in intact animals and in vitro. We have studied the rate of protein degradation by measuring in vitro the release of acid-soluble radioactivity from rat liver slices and tested some factors that control the process. We found a rate of protein degradation of 6.5, or 2% per hr after 1 or 15 hrs of labelling in vivo during the first 90 min. These results indicate that a correlation exists between the rate of production of acid-soluble radioactivity by liver slices and the fast-or slow-turnover rate of the liver proteins. Cyanide and fluoride greatly inhibit the production of acid-soluble radioactivity from both slow- and fast-turnover proteins. Glucagon increases this production while insulin shows an opposite effect. Our preliminary investigations show that liver slices are a suitable surviving medium to study protein catabolism and its modifications under physiological and pathological stimuli.

Animals↗