The best possible lower bound for the Perron root using traces

AbstractLet A be an n×n matrix with real eigenvalues. Wolkowicz and Styan presented bounds for the eigenvalues, using only n, trA, and trA2. We show that their lower bound for the largest eigenvalue works also as a lower bound for the Perron root of A if A is nonnegative and its eigenvalues are not necessarily real. We also show that this bound is optimal under certain conditions. Finally, we solve completely the problem to find the optimal lower bound for the Perron root using only n, trA, and trA2

Analyse fonctionnelle de protéines métal- ou redox- dépendantes chez les plantes

The presence of reactive cysteines confers redox properties and/or the ability to bind metal ions to numerous proteins. This project, organized in several axes, aimed at characterizing proteins with a conserved CxxC motif and possibly a thioredoxin (TRX) fold in plants. It appears that the mitochondrial TRX o1 and o2, the atypical protein disulfide isomerase PDI-A and the chloroplastic glutaredoxin (GRX) S16 from A. thaliana expressed as recombinant proteins in Escherichia coli all incorporated an Fe-S center within homodimers whose function remains to be determined. Analysis of the redox properties of apo-proteins indicates that PDI-A and GRXS16 have little or no oxidoreductase activity respectively although intramolecular disulfide bridges are formed between conserved cysteines. In the case of GRXS16, its redox state would be regulated by light as the disulfide bridge is reducible by TRX but not by glutathione. The last research axis concerned the study of the properties of the MIA40 oxidoreductase and the ERV1 flavine oxidase, involved in the import and oxidative folding of proteins within the inter-membrane space of mitochondria. The results suggest that the singularity of this system in plants is based on the atypical structure of ERV1 and its ability to oxidize proteins in the presence of glutathione but in the absence of MIA40, which is essential in yeast or humans.La présence de cystéines réactives confère à de nombreuses protéines des propriétés redox et/ou la capacité à lier des ions métalliques. Ce projet, organisé en plusieurs axes, visait à caractériser des protéines possédant un motif CxxC conservé et éventuellement un repliement de type thiorédoxine (TRX) chez les plantes. Il s’avère que les TRX o1 et o2 mitochondriales, la protéine disulfure isomérase atypique PDI-A et la glutarédoxine (GRX) S16 chloroplastique d’A. thaliana exprimées sous formes recombinantes dans Escherichia coli incorporent toutes un centre Fe-S au sein d’homodimères dont la fonction reste à déterminer. L’analyse des propriétés redox des apo-protéines indiquent que la PDI–A et la GRXS16 ne possèdent pas ou peu d’activité oxydoréductase respectivement bien que des ponts disulfure intramoléculaires soient formés entre cystéines conservées. Dans le cas de la GRXS16, la régulation de son état redox se ferait via la lumière puisque le pont disulfure est réductible par des TRX mais pas par le glutathion. Le dernier axe de recherche concernait l’étude des propriétés de l’oxydoréductase MIA40 et de la flavine oxydase ERV1, impliquées dans l’import et le repliement oxydatif de protéines au sein de l’espace inter-membranaire des mitochondries. Les résultats suggèrent que la singularité de ce système chez les plantes repose sur la structure atypique d’ERV1 et sa capacité à oxyder des protéines en présence de glutathion mais en absence de MIA40, qui est en revanche indispensable chez la levure ou l’homme

Functional analysis of metal- or redox- dependent proteins in plants

La présence de cystéines réactives confère à de nombreuses protéines des propriétés redox et/ou la capacité à lier des ions métalliques. Ce projet, organisé en plusieurs axes, visait à caractériser des protéines possédant un motif CxxC conservé et éventuellement un repliement de type thiorédoxine (TRX) chez les plantes. Il s’avère que les TRX o1 et o2 mitochondriales, la protéine disulfure isomérase atypique PDI-A et la glutarédoxine (GRX) S16 chloroplastique d’A. thaliana exprimées sous formes recombinantes dans Escherichia coli incorporent toutes un centre Fe-S au sein d’homodimères dont la fonction reste à déterminer. L’analyse des propriétés redox des apo-protéines indiquent que la PDI–A et la GRXS16 ne possèdent pas ou peu d’activité oxydoréductase respectivement bien que des ponts disulfure intramoléculaires soient formés entre cystéines conservées. Dans le cas de la GRXS16, la régulation de son état redox se ferait via la lumière puisque le pont disulfure est réductible par des TRX mais pas par le glutathion. Le dernier axe de recherche concernait l’étude des propriétés de l’oxydoréductase MIA40 et de la flavine oxydase ERV1, impliquées dans l’import et le repliement oxydatif de protéines au sein de l’espace intermembranaire des mitochondries. Les résultats suggèrent que la singularité de ce système chez les plantes repose sur la structure atypique d’ERV1 et sa capacité à oxyder des protéines en présence de glutathion mais en absence de MIA40, qui est en revanche indispensable chez la levure ou l’homme.The presence of reactive cysteines confers redox properties and/or the ability to bind metal ions to numerous proteins. This project, organized in several axes, aimed at characterizing proteins with a conserved CxxC motif and possibly a thioredoxin (TRX) fold in plants. It appears that the mitochondrial TRX o1 and o2, the atypical protein disulfide isomerase PDI-A and the chloroplastic glutaredoxin (GRX) S16 from A. thaliana expressed as recombinant proteins in Escherichia coli all incorporated an Fe-S center within homodimers whose function remains to be determined. Analysis of the redox properties of apo-proteins indicates that PDI-A and GRXS16 have little or no oxidoreductase activity respectively although intramolecular disulfide bridges are formed between conserved cysteines. In the case of GRXS16, its redox state would be regulated by light as the disulfide bridge is reducible by TRX but not by glutathione. The last research axis concerned the study of the properties of the MIA40 oxidoreductase and the ERV1 flavine oxidase, involved in the import and oxidative folding of proteins within the inter-membrane space of mitochondria. The results suggest that the singularity of this system in plants is based on the atypical structure of ERV1 and its ability to oxidize proteins in the presence of glutathione but in the absence of MIA40, which is essential in yeast or humans

Structural Insights into a Fusion Protein between a Glutaredoxin-like and a Ferredoxin-Disulfide Reductase Domain from an Extremophile Bacterium

In eukaryotic photosynthetic organisms, ferredoxin–thioredoxin reductases (FTRs) are key proteins reducing several types of chloroplastic thioredoxins (TRXs) in light conditions. The electron cascade necessary to reduce oxidized TRXs involves a pair of catalytic cysteines and a [4Fe–4S] cluster present at the level of the FTR catalytic subunit, the iron–sulfur cluster receiving electrons from ferredoxins. Genomic analyses revealed the existence of FTR orthologs in non-photosynthetic organisms, including bacteria and archaea, referred to as ferredoxin-disulfide reductase (FDR) as they reduce various types of redoxins. In this study, we describe the tridimensional structure of a natural hybrid protein formed by an N-terminal glutaredoxin-like domain fused to a FDR domain present in the marine bacterium Desulfotalea psychrophila Lsv54. This structure provides information on how and why the absence of the variable subunit present in FTR heterodimer which normally protects the Fe–S cluster is dispensable in FDR proteins. In addition, modelling of a tripartite complex based on the existing structure of a rubredoxin (RBX)–FDR fusion present in anaerobic methanogen archaea allows recapitulating the electron flow involving these RBX, FDR and GRX protein domains

MIA40 of Arabidopsis thaliana is essential for the optimal in vitro oxidation of TIM and COX proteins

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Oxidation of <i>Arabidopsis thaliana</i> COX19 Using the Combined Action of ERV1 and Glutathione

Protein import and oxidative folding within the intermembrane space (IMS) of mitochondria relies on the MIA40–ERV1 couple. The MIA40 oxidoreductase usually performs substrate recognition and oxidation and is then regenerated by the FAD-dependent oxidase ERV1. In most eukaryotes, both proteins are essential; however, MIA40 is dispensable in Arabidopsis thaliana. Previous complementation experiments have studied yeast mia40 mutants expressing a redox inactive, but import-competent versions of yeast Mia40 using A. thaliana ERV1 (AtERV1) suggest that AtERV1 catalyzes the oxidation of MIA40 substrates. We assessed the ability of both yeast and Arabidopsis MIA40 and ERV1 recombinant proteins to oxidize the apo-cytochrome reductase CCMH and the cytochrome c oxidase assembly protein COX19, a typical MIA40 substrate, in the presence or absence of glutathione, using in vitro cysteine alkylation and cytochrome c reduction assays. The presence of glutathione used at a physiological concentration and redox potential was sufficient to support the oxidation of COX19 by AtERV1, providing a likely explanation for why MIA40 is not essential for the import and oxidative folding of IMS-located proteins in Arabidopsis. The results point to fundamental biochemical differences between Arabidopsis and yeast ERV1 in catalyzing protein oxidation

Structural Insights into a Fusion Protein between a Glutaredoxin-like and a Ferredoxin-Disulfide Reductase Domain from an Extremophile Bacterium

In eukaryotic photosynthetic organisms, ferredoxin–thioredoxin reductases (FTRs) are key proteins reducing several types of chloroplastic thioredoxins (TRXs) in light conditions. The electron cascade necessary to reduce oxidized TRXs involves a pair of catalytic cysteines and a [4Fe–4S] cluster present at the level of the FTR catalytic subunit, the iron–sulfur cluster receiving electrons from ferredoxins. Genomic analyses revealed the existence of FTR orthologs in non-photosynthetic organisms, including bacteria and archaea, referred to as ferredoxin-disulfide reductase (FDR) as they reduce various types of redoxins. In this study, we describe the tridimensional structure of a natural hybrid protein formed by an N-terminal glutaredoxin-like domain fused to a FDR domain present in the marine bacterium Desulfotalea psychrophila Lsv54. This structure provides information on how and why the absence of the variable subunit present in FTR heterodimer which normally protects the Fe–S cluster is dispensable in FDR proteins. In addition, modelling of a tripartite complex based on the existing structure of a rubredoxin (RBX)–FDR fusion present in anaerobic methanogen archaea allows recapitulating the electron flow involving these RBX, FDR and GRX protein domains

Atypical protein disulfide isomerases (PDI) : Comparison of the molecular and catalytic properties of poplar PDI-A and PDI-M with PDI-L1A

Protein disulfide isomerases are overwhelmingly multi-modular redox catalysts able to perform the formation, reduction or isomerisation of disulfide bonds.[br/] We present here the biochemical characterization of three different poplar PDI isoforms. PDI-A is characterized by a single catalytic Trx module, the so-called a domain, whereas PDI-L1a and PDI-M display an a-b-b’-a’ and a°-a-b organisation respectively. Their activities have been tested in vitro using purified recombinant proteins and a series of model substrates as insulin, NADPH thioredoxin reductase, NADP malate dehydrogenase (NADP-MDH), peroxiredoxins or RNase A.[br/] We demonstrated that PDI-A exhibited none of the usually reported activities, although the cysteines of the WCKHC active site signature are able to form a disulfide with a redox midpoint potential of -170 mV at pH 7.0. The fact that it is able to bind a [Fe2S2] cluster upon Escherichia coli expression and anaerobic purification might indicate that it does not have a function in dithiol-disulfide exchange reactions. The two other proteins were able to catalyze oxidation or reduction reactions, PDI-L1a being more efficient in most cases, except that it was unable to activate the non-physiological substrate NADP-MDH, in contrast to PDI-M. To further evaluate the contribution of the catalytic domains of PDI-M, the dicysteinic motifs have been independently mutated in each a domain.[br/] The results indicated that the two a domains seem interconnected and that the a° module preferentially catalyzed oxidation reactions whereas the a module catalyzed reduction reactions, in line with the respective redox potentials of -170 mV and -190 mV at pH 7.0. Overall, these in vitro results illustrate that the number and position of a and b domains influence the redox properties and substrate recognition (both electron donors and acceptors) of PDI which contributes to understand why this protein family expanded along evolution

Towards the function of the chloroplastic Arabidopsis thaliana glutaredoxin S16

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