Biomedical subjects

Marzia Arese

Publications and source records attributed to Marzia Arese.

8 recordsLinked to original sources

PubMed2026

Serine: From Metabolic Intermediate to Signaling Entity.

Serine, a nonessential amino acid classically defined as a precursor for protein synthesis and one-carbon metabolism, is increasingly recognized as a signaling metabolite that links the cellular metabolic status to regulatory decision-making. Intracellular serine availability is shaped by nutrient conditions, glycolytic flux, and activity of the serine synthesis pathway, and these fluctuations are sensed to elicit coordinated metabolic and signaling responses. This review discusses mechanisms by which serine modulates cell growth and stress responses, with particular emphasis on its interaction with central nutrient-sensing pathways, including mTORC1 and the integrated stress response. In parallel, serine-driven one-carbon metabolism is examined for its role in supporting nucleotide biosynthesis, methylation reactions, and redox homeostasis through folate-dependent pathways and NADPH generation, thereby coupling anabolic processes to the maintenance of redox balance and genome integrity. In addition to intracellular functions, serine contributes to intercellular signaling. Conversion of l-serine to d-serine mediates neuromodulatory activity via N-methyl-d-aspartate receptors, while serine availability also influences immune cell function, inflammatory signaling, and host-microbe interactions. Dysregulation of serine metabolism and signaling is further considered in the context of disease states, including cancer, neurodegeneration, and metabolic disorders. Together, these observations support a framework in which serine functions as an information-bearing metabolic signal that coordinates the biosynthetic capacity with cellular adaptation and intercellular communication.

Serine↗

PubMed2006

Nitric oxide and the respiratory enzyme.

Available information on the molecular mechanisms by which nitric oxide (NO) controls the activity of the respiratory enzyme (cytochrome-c-oxidase) is reviewed. We report that, depending on absolute electron flux, NO at physiological concentrations reversibly inhibits cytochrome-c-oxidase by two alternative reaction pathways, yielding either a nitrosyl- or a nitrite-heme a3 derivative. We address a number of hypotheses, envisaging physiological and/or pathological effects of the reactions between NO and cytochrome-c-oxidase.

Animals↗

PubMed2003

Expression of a fully functional cd1 nitrite reductase from Pseudomonas aeruginosa in Pseudomonas stutzeri.

Nitrite reductases are redox enzymes catalysing the one electron reduction of nitrite to nitrogen monoxide (NO) within the bacterial denitrification process. We have cloned the gene for cd(1) nitrite reductase (Pa-nirS) from Pseudomonas aeruginosa into the NiRS(-) strain MK202 of Pseudomonas stutzeri and expressed the enzyme under denitrifying conditions. In the MK202 strain, denitrification is abolished by the disruption of the endogenous nitrite reductase gene; thus, cells can be grown only in the presence of oxygen. After complementation with Pa-nirS gene, cells supplemented with nitrate can be grown in the absence of oxygen. The presence of nitrite reductase was proven in vivo by the demonstration of NO production, showing that the enzyme was expressed in the active form, containing both heme c and d(1). A purification procedure for the recombinant PaNir has been developed, based on the P. aeruginosa purification protocol; spectroscopic analysis of the purified protein fully confirms the presence of the d(1) heme cofactor. Moreover, the functional characterisation of the recombinant NiR has been carried out by monitoring the production of NO by the purified NiR enzyme in the presence of nitrite by an NO electrode. The full recovery of the denitrification properties in the P. stutzeri MK202 strain by genetic complementation with Pa-NiR underlines the high homology between enzymes of nitrogen oxianion respiration. Our work provides an expression system for cd(1) nitrite reductase and its site-directed mutants in a non-pathogenic strain and is a starting point for the in vivo study of recombinant enzyme variants.

Cloning, Molecular↗

PubMed2003

The molecular mechanisms by which nitric oxide controls mitochondrial complex IV.

This mini-review of focussed on the information available on the molecular mechanisms by which NO controls the function of mitochondrial cytochrome c oxidase and thereby cell respiration. The reaction mechanisms are described as dissected in vitro and recently confirmed in cell cultures, whereby two reaction pathways have been identified, leading to accumulation of either the [a3(2+)NO]-nitrosyl or the [a3(3+)NO2-]-nitrite derivative of the enzyme. The experimental data and the theoretical computation analysis, supporting the hypothesis that one pathway prevails on the other depending on the electron flow level through the respiratory chain, are discussed. Finally, the patho-physiological implications of the reaction between NO and CcOX have been also outlined.

Animals↗

PubMed2002

Cytochrome bo(3) from Escherichia coli: the binding and turnover of nitric oxide.

The reaction of nitric oxide (NO) with fast and reduced cytochrome bo(3)(cyt bo(3)) from Escherichia coli has been investigated. The stoichiometry of NO binding to cyt bo(3) was determined using an NO electrode in the [NO] range 1-14 microM. Under reducing conditions, the initial decrease in [NO] following the addition of cyt bo(3) corresponded to binding of 1 NO molecule per cyt bo(3) functional unit. After this "rapid" NO binding phase, there was a slow, but significant rate of NO consumption ( approximately 0.3molNOmol bo(3)(-1)min(-1)), indicating that cyt bo(3) possesses a low level of NO reductase activity. The binding of NO to fast pulsed enzyme was also investigated. The results show that in the [NO] range used (1-14 microM) both fast and pulsed oxidised cyt bo(3) bind NO with a stoichiometry of 1:1 with an observed dissociation constant of K(d)=5.6+/-0.6 microM and that NO binding was inhibited by the presence of Cl(-). The binding of nitrite to the binuclear centre causes spectral changes similar to those observed upon NO binding to fast cyt bo(3). These results are discussed in relation to the model proposed by Wilson and co-workers [FEBS Lett. 414 (1997) 281] where the binding of NO to Cu(B)(II) results in the formation of the nitrosonium (Cu(B)(I)-NO(+)) complex. NO(+) then reacts with OH(-), a Cu(B) ligand, to form nitrite, which can bind at the binuclear centre. This work suggests for the first time that the binding of NO to oxidised cyt bo(3) does result in the reduction of Cu(B).

Binding Sites↗

PubMed2002

Cyanide binding to cd(1) nitrite reductase from Pseudomonas aeruginosa: role of the active-site His369 in ligand stabilization.

Cyanide binding to fully reduced Pseudomonas aeruginosa cd(1) nitrite reductase (Pa cd(1) NiR) has been investigated for the wild-type enzyme and a site-directed mutant in which the active-site His369 was replaced by Ala. This mutation reduces the affinity toward cyanide (by approximately 13-fold) and especially decreases the rate of binding of cyanide to the reduced d(1) heme (by approximately 100-fold). The crystal structure of wild-type reduced Pa cd(1) NiR saturated with cyanide was determined to a resolution of 2.7 A. Cyanide binds to the iron of the d(1) heme, with an Fe-C-N angle of 168 degrees for both subunits of the dimer and only His369 is within hydrogen bonding distance of the nitrogen atom of the ligand. These results suggest that in Pa cd(1) NiR the invariant distal residue His369 plays a dominant role in controlling the binding of anionic ligands and allow the discussion of the mechanism of cyanide binding to the wild-type enzyme.

Amino Acid Substitution↗

PubMed2002

Pseudomonas aeruginosa cytochrome C(551): probing the role of the hydrophobic patch in electron transfer.

Cytochrome c(551) from Pseudomonas aeruginosa is a monomeric redox protein of 82 amino-acid residues, involved in dissimilative denitrification as the physiological electron donor of cd(1) nitrite reductase. The distribution of charged residues on the surface of c(551) is very anisotropic: one side is richer in acidic residues whereas the other shows a ring of positive side chains, mainly lysines, located at the border of an hydrophobic patch which surrounds the heme crevice. In order to map in cytochrome c(551) the surface involved in electron transfer, we have introduced specific mutations in three residues belonging to the hydrophobic patch, namely Val23-->Asp, Pro58-->Ala and Ile59-->Glu. The effect of these mutations was analyzed studying both the self-exchange rate and the electron-transfer activity towards P. aeruginosa cd(1) nitrite reductase, the physiological partner and P. aeruginosa azurin, a copper protein often used as a model redox partner in vitro. Our results show that introduction of a negative charge in the hydrophobic patch severely hampers both homonuclear and heteronuclear electron transfer.

Azurin↗

PubMedDate not supplied

Nitric oxide and mitochondrial complex IV.

Micromolar nitric oxide (NO) rapidly (ms) inhibits cytochrome c oxidase in turnover with physiological substrates. Two reaction mechanisms have been identified leading, respectively, to formation of a nitrosyl- [a3(2+) -NO] or a nitrite- [a3(3+) -NO2-] derivative of the enzyme. In the presence of O2, the nitrosyl adduct recovers activity slowly, following NO displacement at k' approximately equal to 0.01 s(-1) (37 degrees C); the recovery of the nitrite adduct is much faster. Relevant to pathophysiology, the enzyme does not degrade NO by following the first mechanism, whereas by following the second one it promotes NO oxidation and disposal as nitrite/nitrate. The reaction between NO and cytochrome c oxidase has been investigated at different integration levels of the enzyme, including the in situ state, such as in mouse liver mitochondria or cultured human SY5Y neuroblastoma cells. The respiratory chain is inhibited by NO, either supplied exogenously or produced endogenously via the NO synthase activation. Inhibition of respiration is reversible, although it remains to be clarified whether reversibility is always full and how it depends on concentration of and time of exposure to NO. Oxygraphic measurements show that cultured cells or isolated state 4 mitochondria exposed to micromolar (or less) NO recover from NO inhibition rapidly, as if the nitrite reaction was predominant. Mitochondria in state 3 display a slightly more persistent inhibition than in state 4, possibly due to a higher accumulation of the nitrosyl adduct. Among a number of parameters that appear to control the switch over between the two mechanisms, the concentration of reductants (reduced cytochrome c) at the cytochrome c oxidase site has been proved to be the most relevant one.

Animals↗