RESUMO

Bacterial and fungal large-size subunit catalases (LSCs) are like small-size subunit catalases (SSCs) but have an additional C-terminal domain (CT). The catalytic domain is conserved at both primary sequence and structural levels and its amino acid composition is optimized to select H2O2 over water. The CT is structurally conserved, has an amino acid composition similar to very stable proteins, confers high stability to LSCs, and has independent molecular chaperone activity. While heat and denaturing agents increased Neurospora crassa catalase-1 (CAT-1) activity, a CAT-1 version lacking the CT (C63) was no longer activated by these agents. The addition of catalase-3 (CAT-3) CT to the CAT-1 or CAT-3 catalase domains prevented their heat denaturation in vitro. Protein structural alignments indicated CT similarity with members of the DJ-1/PfpI superfamily and the CT dimers present in LSCs constitute a new type of symmetric dimer within this superfamily. However, only the bacterial Hsp31 proteins show sequence similarity to the bacterial and fungal catalase mobile coil (MC) and are phylogenetically related to MC_CT sequences. LSCs might have originated by fusion of SSC and Hsp31 encoding genes during early bacterial diversification, conferring at the same time great stability and molecular chaperone activity to the novel catalases.

RESUMO

Sporothrix schenckii is a dimorphic fungus, pathogenic to humans and animals, which is usually infective in the yeast form. Reactive oxygen species (ROS) play an important role in the host's defense, damaging the pathogen's DNA, proteins, and lipids. To prevent oxidative damage, the ROS are detoxified by pathogen-derived antioxidant enzymes such as catalases (CATs). In this work, we analyzed the activity and expression level of three S. schenckii genes, designated as CAT1, CAT2, and CAT3, that putatively encoded for three isoforms of monofunctional CAT with a predicted molecular weight of 57.6, 56.2, and 81.4 kDa, respectively. Our results demonstrate that oxidative stress induced by exogenous H2O2 leads to an altered lipid peroxidation, modifying CAT activity and the expression levels of the CAT genes, being CAT1 and CAT3 the genes with the highest expression in response to the oxidizing agent. These results show that CAT isoforms in S. schenckii can be regulated in response to oxidative stress and might help to control ROS homeostasis in the fungus-host interaction.

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Heme catalases remove hydrogen peroxide by catalyzing its dismutation into water and molecular oxygen, thereby protecting the cell from oxidative damage. The Atacama plateau in northern Argentina, located 4000 m above sea level, is a desert area characterized by extreme UV radiation, high salinity and a large temperature variation between day and night. Here, the heme catalase KatE1 from an Atacama Acinetobacter sp. isolate was cloned, expressed and purified, with the aim of investigating its extremophilic properties. Kinetic and stability assays indicate that KatE1 is maximally active at 50°C in alkaline media, with a nearly unchanged specific activity between 0°C and 40°C in the pH range 5.5-11.0. In addition, its three-dimensional crystallographic structure was solved, revealing minimal structural differences compared with its mesophilic and thermophilic analogues, except for a conserved methionine residue on the distal heme side, which is proposed to comprise a molecular adaptation to oxidative damage.

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Dinuclear manganese (II- III) compounds, which are potential models of the active center of catalase, were synthetized. This type of metalloenzymes presents biological importance due to three factors: they are redox catalyst centres, they are able to carry out hydrolytic reactions and they participate in activated processes via Lewis acids. Structurally, their active centre is composed by dinuclear manganese compounds, linked to nitrogen and oxygen donor atoms. An octahedral geometry around the metal ions were found, with acetate, hydroxy and aquo ligands; which can work as molecule bridges between them. The acid medium favours the electronic transfer between Mn3+ - Mn2+ as redox centre at 1.559 V and the consequent oxidation of hydrogen peroxide or organic molecules. The work also reports the data of two chiral novel compounds, [Mn2(S,S(+)Hcpse)4(NaClO4)2(NaOH)(CH4O)]n·[(C2H6O)2]n·[(CH4O)2]n and its respective enantioisomer, in which µ-oxo being as bridge metal centre. The X-ray structural was obtained as well as the optical and magnetic properties using Circular Dichroism, Electronic Paramagnetic Resonance, Magnetic Susceptibility and X-ray photoelectron spectroscopy.

RESUMO

Dimorphic human pathogenic fungi interact with host effector cells resisting their microbicidal mechanisms. Yeast cells are able of surviving within the tough environment of the phagolysosome by expressing an antioxidant defense system that provides protection against host-derived reactive oxygen species (ROS). This includes the production of catalases (CATs). Here we identified and analyzed the role of CAT isoforms in Paracoccidioides, the etiological agent of paracoccidioidomycosis. Firstly, we found that one of these isoforms was absent in the closely related dimorphic pathogen Coccidioides and dermatophytes, but all of them were conserved in Paracoccidioides, Histoplasma and Blastomyces species. We probed the contribution of CATs in Paracoccidioides by determining the gene expression levels of each isoform through quantitative RT-qPCR, in both the yeast and mycelia phases, and during the morphological switch (transition and germination), as well as in response to oxidative agents and during interaction with neutrophils. PbCATP was preferentially expressed in the pathogenic yeast phase, and was associated to the response against exogenous H2O2. Therefore, we created and analyzed the virulence defects of a knockdown strain for this isoform, and found that CATP protects yeast cells from H2O2 generated in vitro and is relevant during lung infection. On the other hand, CATA and CATB seem to contribute to ROS homeostasis in Paracoccidioides cells, during endogenous oxidative stress. CAT isoforms in Paracoccidioides might be coordinately regulated during development and dimorphism, and differentially expressed in response to different stresses to control ROS homeostasis during the infectious process, contributing to the virulence of Paracoccidioides.

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BACKGROUND: In addition to high production levels of conidia, the success of entomopathogenic fungi as biological control agents depends both on their prevalence under the environmental conditions found in open fields (resistance to stress) and on the capacity of these conidia to infect pests. This study compares conidium production, infectivity and resistance to thermal and osmotic stress in two strains of Isaria fumosorosea (ARSEF 3302 and CNRCB1) grown either under a normal atmosphere (21% O2) or using enriched oxygen pulses (26% O2). RESULTS: After 180 h, the ARSEF 3302 strain with 26% O2 pulses increased conidium production nearly fivefold compared with the normal atmosphere, while conidium production by the CNRCB1 strain decreased by 50% under O2 pulses, relative to the values measured with the normal atmosphere. The conidia obtained with 26% O2 pulses had a greater germination rate and resistance to thermal and osmotic stress, in addition to improved infectivity against Galleria mellonella (Lepidoptera) larvae. These findings were associated with an increase in catalase activities for both strains. CONCLUSION: An enriched oxygen atmosphere increases the quality of conidia of both strains of I. fumosorosea, with a variable effect on conidium production.

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