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A Cozzi

Publications and source records attributed to A Cozzi.

49 records · Page 3Linked to original sources
PubMed1989

Expression and structural and functional properties of human ferritin L-chain from Escherichia coli.

The human ferritin L-chain cDNA was cloned into a vector for overproduction in Escherichia coli, under the regulation of a lambda promoter. The plasmid obtained contains the full L-chain coding region modified at the first two codons. It is able to direct the synthesis of the L-chain which can constitute up to 15% of the total soluble protein of bacterial extract. The L-chains assemble to form a ferritin homopolymer with electrophoretic mobility, molecular weight, thermal stability, spectroscopic, and immunological properties analogous to natural ferritin from human liver (95% L-chain). This recombinant L-ferritin is able to incorporate and retain iron in solution at physiological pH values. At variance with the H-ferritin, the L form does not uptake iron at acidic pH values and does not show detectable ferroxidase activity. It is concluded that ferritin L-chain lacks the ferroxidase site present in the H-chain and that the two chains may have specialized functions in intracellular iron metabolism.

Blotting, Western↗
PubMed1988

Mechanism of ferritin iron uptake: activity of the H-chain and deletion mapping of the ferro-oxidase site. A study of iron uptake and ferro-oxidase activity of human liver, recombinant H-chain ferritins, and of two H-chain deletion mutants.

To study the functional differences between human ferritin H- and L-chains and the role of the protein shell in the formation and growth of the ferritin iron core, we have compared the kinetics of iron oxidation and uptake of ferritin purified from human liver (90% L) and of the H-chain homopolymer overproduced in Escherichia coli (100% H). As a control for iron autocatalytic activity, we analyzed the effect of Fe(III) on the iron uptake reaction. The results show that the H-chain homopolymer has faster rates of iron uptake and iron oxidation than liver ferritin in all the conditions analyzed and that the difference is reduced in the conditions in which iron autocatalysis in high: i.e. at pH 7 and in presence of iron core. We have also analyzed the properties of two engineered H-chains, one lacking the last 22 amino acids at the carboxyl terminus and the other missing the first 13 residues at the amino terminus. These mutant proteins assemble in ferritin-like proteins and maintain the ability to catalyze iron oxidation. The deletion at the carboxyl terminus, however, prevents the formation of a stable iron core. It is concluded that the ferritin H-chain has an iron oxidation site which is separated from the sites of iron transfer and hydrolysis and that either the integrity of the molecule or the presence of the amino acid sequences forming the hydrophobic channel is necessary for iron core formation.

Chromosome Deletion↗
PubMed1988

Mechanisms of regulation of ferritin synthesis in rat liver during experimental inflammation.

Liver slices of turpentine-treated rats were incubated in vitro and used as a model to study synthesis and secretion of proteins during the acute-phase response. The synthesis and secretion of typical acute-phase proteins increased after treatment. Similarly, ferritin increased at 24-48 hr after treatment. Serum ferritin showed a slight and transient increase at 6 hr; however, no ferritin was detectable in liver slices medium, indicating no or negligible secretion by this tissue. Northern blot analysis of RNA extracted from total liver homogenate and from free and membrane-bound polyribosomes revealed that turpentine treatment stimulates ferritin synthesis at the translational level, possibly increasing the amount of ferritin mRNA on membrane-bound polysomes.

Acute-Phase Proteins↗
PubMed1988

Characteristics and expression of binding sites specific for ferritin H-chain on human cell lines.

Purified recombinant human ferritin composed solely of H subunit was radiolabeled and incubated with proerythroleukemic K562 human cells. A specific binding was detected, and it could be displaced only by ferritins, natural or recombinant, containing large proportion of the H subunit. The specific ferritin H-chain binding was saturable, and cells showed 17,000 to 23,000 binding sites per cell. The affinity constant measured at 37 degrees C was of 3 x 10(8) M-1. Treatment with pronase eliminated the specific binding. The binding sites were expressed in a high number during the cellular exponential phase of growth and progressively decreased to disappear when cells reached the plateau phase. Treatment of the cells with desferrioxamine increased recombinant H-ferritin binding, while iron had little effect. K562 cells induced to differentiate by hemin failed to bind ferritin H. Ferritin H-chain binding capacity is present on various cell lines such as HL60, lung cancer, and hepatoma cells. Analysis of the binding sites by western blotting showed a peptide with apparent mol wt of about 100 kd.

Binding Sites↗
PubMed1987

Human serum ferritin G-peptide is recognized by anti-L ferritin subunit antibodies and concanavalin-A.

Ferritin was purified from serum of patients with idiopathic haemochromatosis. Analysis on SDS electrophoresis showed that it is composed of two major bands of 19,000 and 23,000 Mr. The smaller peptide has an electrophoretic mobility and immunochemical reactivity similar to that of tissue L subunit. The larger, previously named G subunit, is recognized by concanavalin-A and by anti ferritin L-subunit, but not by anti-H, monoclonal antibodies. All of the antibodies show higher affinity for the L than for the G subunit. Therefore, the G chain appears immunochemically similar, but not identical, to ferritin L chain, and is responsible for serum ferritin binding concanavalin-A.

Antibodies↗
PubMed1987

Characteristics of a ferritin-binding protein present in human serum.

The ferritin present in human serum differs from the ferritins found in tissues and other body fluids in having negligible proportions of H subunits. This has been related to the possible presence of binding factors which would form complexes with H-subunit containing ferritins and thereby determine their rapid clearance and/or interference with immunoassays ('serum inhibition'). In this work we have tried to identify and characterize these binding factors. Dotting and blotting experiments demonstrated an interaction between tissue ferritins and human serum. This was stronger with human heart and recombinant H-type ferritin obtained from E. coli than with human liver ferritin. The serum binder appeared to be a glycoprotein migrating in the beta-2 region and with a molecular weight of about 200,000 and pI between 4 and 5. Two different approaches to purification of the ferritin-binding protein yielded enriched fractions containing also the complement proteins C3 and C4, the plasma protease inhibitor alpha-2-macroglobulin, and immunoglobulins. These in vitro findings may have physiological relevance.

Blood Proteins↗
PubMed1983

Chronobiological aspects of headache syndromes due to sellar or pituitary pathology.

A chronobiological study was carried out in headache syndromes due to empty sella or to pituitary G.H.- and PRL-secreting adenomas. In the empty sella syndrome only the chrono-organization of G.H. secretion was disturbed, whereas pl. PRL exhibited the usual circadian pattern. The circadian rhythms of pl. G.H. and pl. PRL were abolished in G.H.- and PRL-secreting pituitary tumors, respectively, and were again detectable when patients were cured by selective transsphenoidal adenomectomy. A normal circadian rhythmicity of pl. cortisol was demonstrable in the empty sella syndrome and in pituitary adenomas, both before and after surgery.

Adenoma↗
PubMed1983

[Pharmacokinetics of amiodarone in one case of acute oral intoxication].

A wealth of data about the clinical use and the therapeutic efficacy of chronic treatment with amiodarone has been reported, while the acute effects of this agent are less known. In one case of acute oral intoxication with 8 g of amiodarone for suicidal purposes, we have investigated the pharmacokinetics of amiodarone, the thyroid function and the variations of some clinical parameters, such as heart rate and QT interval. Other possible side-effects have been looked for. We have not observed substantial pharmacokinetic differences between our case and studies carried out after chronic oral or intravenous administration. Unlike chronic administration, the acute oral load was not followed by the appearance of toxic effects. The absence of toxic phenomena can be explained by the poor bioavailability of amiodarone, which is known to require long periods for a complete distribution to the tissues and target organs.

Adult↗