Lysis of Paracoccidioides brasiliensis by Zygosporium geminatum

San-Blas, Gioconda; Moreno, Belisario; Calcagno, A. M.; San-Blas, Felipe

doi:10.1046/j.1365-280x.1998.00121.x

Cited by 2 publications

(2 citation statements)

References 23 publications

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“…(1 → 3)‐α‐Glucanases are glycoside hydrolases that catalyze the degradation of (1 → 3)‐α‐glucan, a water‐insoluble polysaccharide in the cell wall of many fungi. Both bacteria [1–5] and fungi [6–16] are known to produce (1 → 3)‐α‐glucanases, but their respective glucanases are quite distinct in amino acid sequence. Based on amino acid sequence similarities, the fungal (1 → 3)‐α‐glucanases have been assigned a separate family of glycoside hydrolases, GH‐71.…”

Section: Introductionmentioning

confidence: 99%

Mechanism of action of the endo‐(1 → 3)‐α‐glucanase MutAp from the mycoparasitic fungus Trichoderma harzianum

et al. 2006

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(1-->3)-alpha-glucanases catalyze the hydrolysis of fungal cell wall (1-->3)-alpha-glucan, and function during cell division of yeasts containing this cell wall component or act in mycoparasitic processes. Here, we characterize the mechanism of action of the (1-->3)-alpha-glucanase MutAp from the mycoparasitic fungus Trichoderma harzianum. We observed that MutAp releases predominantly beta-glucose upon hydrolysis of crystalline (1-->3)-alpha-glucan, indicating inversion of the anomeric configuration. After having identified (1-->3)-alpha-glucan tetrasaccharide as the minimal substrate for MutAp, we showed that reduced (1-->3)-alpha-glucan pentasaccharide is cleaved into a trisaccharide and a reduced disaccharide, demonstrating that MutAp displays endo-hydrolytic activity. We propose a model for the catalytic mechanism of MutAp, whereby the enzyme breaks an intrachain glycosidic linkage of (1-->3)-alpha-glucan, and then continues its hydrolysis towards the non-reducing end by releasing beta-glucose residues in a processive manner.

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Section: Introductionmentioning

confidence: 99%

Mechanism of action of the endo‐(1 → 3)‐α‐glucanase MutAp from the mycoparasitic fungus Trichoderma harzianum

et al. 2006

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“…Reactions were stopped by heating at 100°C [40]. Free reducing ends were analyzed using the colorimetric bicinchoninic acid (BCA) assay [41].…”

Section: Methodsmentioning

confidence: 99%

Biochemical Characterization of Paracoccidioides brasiliensis α-1,3-Glucanase Agn1p, and Its Functionality by Heterologous Expression in Schizosaccharomyces pombe

et al. 2013

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α-1,3-Glucan is present as the outermost layer of the cell wall in the pathogenic yeastlike (Y) form of Paracoccidioides brasiliensis. Based on experimental evidence, this polysaccharide has been proposed as a fungal virulence factor. To degrade α-1,3-glucan and allow remodeling of the cell wall, α-1,3-glucanase is required. Therefore, the study of this enzyme, its encoding gene, and regulatory mechanisms, might be of interest to understand the morphogenesis and virulence process in this fungus. A single gene, orthologous to other fungal α-1,3-glucanase genes, was identified in the Paracoccidioides genome, and labeled AGN1. Transcriptional levels of AGN1 and AGS1 (α-1,3-glucan synthase-encoding gene) increased sharply when the pathogenic Y phase was cultured in the presence of 5% horse serum, a reported booster for cell wall α-1,3-glucan synthesis in this fungus. To study the biochemical properties of P. brasiliensis Agn1p, the enzyme was heterologously overexpressed, purified, and its activity profile determined by means of the degradation of carboxymethyl α-1,3-glucan (SCMG, chemically modified from P. brasiliensis α-1,3-glucan), used as a soluble substrate for the enzymatic reaction. Inhibition assays, thin layer chromatography and enzymatic reactions with alternative substrates (dextran, starch, chitin, laminarin and cellulose), showed that Agn1p displays an endolytic cut pattern and high specificity for SCMG. Complementation of a Schizosaccharomyces pombe agn1Δ strain with the P. brasiliensis AGN1 gene restored the wild type phenotype, indicating functionality of the gene, suggesting a possible role of Agn1p in the remodeling of P. brasiliensis Y phase cell wall. Based on amino acid sequence, P. brasiliensis Agn1p, groups within the family 71 of fungal glycoside hydrolases (GH-71), showing similar biochemical characteristics to other members of this family. Also based on amino acid sequence alignments, we propose a subdivision of fungal GH-71 into at least five groups, for which specific conserved sequences can be identified.

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Lysis of Paracoccidioides brasiliensis by Zygosporium geminatum

Cited by 2 publications

References 23 publications

Mechanism of action of the endo‐(1 → 3)‐α‐glucanase MutAp from the mycoparasitic fungus Trichoderma harzianum

Mechanism of action of the endo‐(1 → 3)‐α‐glucanase MutAp from the mycoparasitic fungus Trichoderma harzianum

Biochemical Characterization of Paracoccidioides brasiliensis α-1,3-Glucanase Agn1p, and Its Functionality by Heterologous Expression in Schizosaccharomyces pombe

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