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1.
PLoS Pathog ; 19(5): e1011393, 2023 May.
Artículo en Inglés | MEDLINE | ID: mdl-37235600

RESUMEN

To gain a better insight of how Copper (Cu) ions toxify cells, metabolomic analyses were performed in S. aureus strains that lacks the described Cu ion detoxification systems (ΔcopBL ΔcopAZ; cop-). Exposure of the cop- strain to Cu(II) resulted in an increase in the concentrations of metabolites utilized to synthesize phosphoribosyl diphosphate (PRPP). PRPP is created using the enzyme phosphoribosylpyrophosphate synthetase (Prs) which catalyzes the interconversion of ATP and ribose 5-phosphate to PRPP and AMP. Supplementing growth medium with metabolites requiring PRPP for synthesis improved growth in the presence of Cu(II). A suppressor screen revealed that a strain with a lesion in the gene coding adenine phosphoribosyltransferase (apt) was more resistant to Cu. Apt catalyzes the conversion of adenine with PRPP to AMP. The apt mutant had an increased pool of adenine suggesting that the PRPP pool was being redirected. Over-production of apt, or alternate enzymes that utilize PRPP, increased sensitivity to Cu(II). Increasing or decreasing expression of prs resulted in decreased and increased sensitivity to growth in the presence of Cu(II), respectively. We demonstrate that Prs is inhibited by Cu ions in vivo and in vitro and that treatment of cells with Cu(II) results in decreased PRPP levels. Lastly, we establish that S. aureus that lacks the ability to remove Cu ions from the cytosol is defective in colonizing the airway in a murine model of acute pneumonia, as well as the skin. The data presented are consistent with a model wherein Cu ions inhibits pentose phosphate pathway function and are used by the immune system to prevent S. aureus infections.


Asunto(s)
Cobre , Staphylococcus aureus , Animales , Ratones , Staphylococcus aureus/metabolismo , Vía de Pentosa Fosfato , Ribosa-Fosfato Pirofosfoquinasa/genética , Ribosa-Fosfato Pirofosfoquinasa/metabolismo , Fosforribosil Pirofosfato/metabolismo , Adenina
2.
Mol Microbiol ; 115(4): 554-573, 2021 04.
Artículo en Inglés | MEDLINE | ID: mdl-33034093

RESUMEN

S. aureus USA300 isolates utilize the copBL and copAZ gene products to prevent Cu intoxication. We created and examined a ΔcopAZ ΔcopBL mutant strain (cop-). The cop- strain was sensitive to Cu and accumulated intracellular Cu. We screened a transposon (Tn) mutant library in the cop- background and isolated strains with Tn insertions in the mntABC operon that permitted growth in the presence of Cu. The mutations were in mntA and they were recessive. Under the growth conditions utilized, MntABC functioned in manganese (Mn) import. When cultured with Cu, strains containing a mntA::Tn accumulated less Cu than the parent strain. Mn(II) supplementation improved growth when cop- was cultured with Cu and this phenotype was dependent upon the presence of MntR, which is a repressor of mntABC transcription. A ΔmntR strain had an increased Cu load and decreased growth in the presence of Cu, which was abrogated by the introduction of mntA::Tn. Over-expression of mntABC increased cellular Cu load and sensitivity to Cu. The presence of a mntA::Tn mutation protected iron-sulfur (FeS) enzymes from inactivation by Cu. The data presented are consistent with a model wherein defective MntABC results in decreased cellular Cu accumulation and protection to FeS enzymes from Cu poisoning.


Asunto(s)
Proteínas de Transporte de Catión/fisiología , Cobre/metabolismo , Cobre/farmacología , Regulación Bacteriana de la Expresión Génica , Manganeso/metabolismo , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/fisiología , Antibacterianos/metabolismo , Antibacterianos/farmacología , Proteínas Bacterianas/fisiología , ADN Bacteriano , Humanos , Hierro/metabolismo , Proteínas Hierro-Azufre/metabolismo , Proteínas de Transporte de Membrana/fisiología , Mutagénesis Insercional , Operón , ARN Bacteriano , Proteínas Represoras/fisiología , Infecciones Estafilocócicas/microbiología
3.
Front Microbiol ; 7: 748, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-27242761

RESUMEN

Members of the genus Leptospirillum are aerobic iron-oxidizing bacteria belonging to the phylum Nitrospira. They are important members of microbial communities that catalyze the biomining of sulfidic ores, thereby solubilizing metal ions. These microorganisms live under extremely acidic and metal-loaded environments and thus must tolerate high concentrations of reactive oxygen species (ROS). Cobalamin (vitamin B12) is a cobalt-containing tetrapyrrole cofactor involved in intramolecular rearrangement reactions and has recently been suggested to be an intracellular antioxidant. In this work, we investigated the effect of the exogenous addition of cobalamin on oxidative stress parameters in Leptospirillum group II strain CF-1. Our results revealed that the external supplementation of cobalamin reduces the levels of intracellular ROSs and the damage to biomolecules, and also stimulates the growth and survival of cells exposed to oxidative stress exerted by ferric ion, hydrogen peroxide, chromate and diamide. Furthermore, exposure of strain CF-1 to oxidative stress elicitors resulted in the transcriptional activation of the cbiA gene encoding CbiA of the cobalamin biosynthetic pathway. Altogether, these data suggest that cobalamin plays an important role in redox protection of Leptospirillum strain CF-1, supporting survival of this microorganism under extremely oxidative environmental conditions. Understanding the mechanisms underlying the protective effect of cobalamin against oxidative stress may help to develop strategies to make biomining processes more effective.

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