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Your search keyword '"Arias, Diego G."' showing total 37 results
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37 results on '"Arias, Diego G."'

36. Functional and structural characterization of an endo-β-1,3-glucanase from Euglena gracilis.

Author

Calloni, Rodrigo D., Muchut, Robertino J., Garay, Alberto S., Arias, Diego G., Iglesias, Alberto A., and Guerrero, Sergio A.

Subjects
*EUGLENA gracilis, *GEL permeation chromatography, *SACCHAROMYCES cerevisiae, *POLYSACCHARIDES, *PROTEIN folding, *ETHANOL, *BETA-glucans
Abstract

Endo-β-1,3-glucanases from several organisms have attracted much attention in recent years because of their capability for in vitro degrading β-1,3-glucan as a critical step for both biofuels production and short-chain oligosaccharides synthesis. In this study, we biochemically characterized a putative endo-β-1,3-glucanase (Egr GH64) belonging to the family GH64 from the single-cell protist Euglena gracilis. The gene coding for the enzyme was heterologously expressed in a prokaryotic expression system supplemented with 3% (v/v) ethanol to optimize the recombinant protein right folding. Thus, the produced enzyme was highly purified by immobilized-metal affinity and gel filtration chromatography. The enzymatic study demonstrated that Egr GH64 could hydrolyze laminarin (K M 23.5 mg ml−1, k cat 1.20 s−1) and also, but with less enzymatic efficiency, paramylon (K M 20.2 mg ml−1, k cat 0.23 ml mg−1 s−1). The major product of the hydrolysis of both substrates was laminaripentaose. The enzyme could also use ramified β-glucan from the baker's yeast cell wall as a substrate (K M 2.10 mg ml−1, k cat 0.88 ml mg−1 s−1). This latter result, combined with interfacial kinetic analysis evidenced a protein's greater efficiency for the yeast polysaccharide, and a higher number of hydrolysis sites in the β-1,3/β-1,6-glucan. Concurrently, the enzyme efficiently inhibited the fungal growth when used at 1.0 mg/mL (15.4 μM). This study contributes to assigning a correct function and determining the enzymatic specificity of Egr GH64, which emerges as a relevant biotechnological tool for processing β-glucans. [Display omitted] • We identified a putative endo-β-1,3 glucanase (EgrGH64) in Euglena gracilis. • The gene was cloned and heterologously expressed in Escherichia coli. • EgrGH64 hydrolyzed paramylon, laminarin, and yeast cell wall. • EgrGH64 probed to be a laminaripentaose-producing enzyme. • The purified enzyme inhibited yeast growth. [ABSTRACT FROM AUTHOR]

Published
2023
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37. Cloning, expression, and characterization of a dithiol glutaredoxin from Trypanosoma cruzi.

Author

Marquez VE, Arias DG, Piattoni CV, Robello C, Iglesias AA, and Guerrero SA

Subjects
Amino Acid Sequence, Cloning, Molecular, Molecular Sequence Data, Oxidation-Reduction, Recombinant Proteins genetics, Recombinant Proteins metabolism, Sequence Alignment, Trypanosoma cruzi genetics, Glutaredoxins genetics, Glutaredoxins metabolism, Trypanosoma cruzi enzymology
Abstract

Glutaredoxins play an important role in cellular functionality. A putative dithiol glutaredoxin is encoded in the genome of Trypanosoma cruzi. We cloned the gene and obtained the recombinant protein, which behaved as a typical thioltransferase. Activity was variable and dependent on the nature of reducer or oxidant agent used, or both. Epimastigote extracts exhibited similar activity, suggesting the occurrence of the protein in the parasite. Results support a redox scenario in T. cruzi, with glutaredoxin being involved mainly in reduction of glutathione disulfide as well as in deglutathionylation of target proteins.

Published
2010
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