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Characterization and crystal structure of a first fungal glyoxylate reductase from Paecilomyes thermophila

Paper ID Volume ID Publish Year Pages File Format Full-Text
16937 42623 2014 8 PDF Available
Title
Characterization and crystal structure of a first fungal glyoxylate reductase from Paecilomyes thermophila
Abstract

•A glyoxylate reductase (PtGR) from a fungus was biochemically and structurally characterized.•PtGR forms a homodimer and has a high specificity toward glyoxylate.•PtGR displayed broad pH stability (pH 4.5–10) and an optimal temperature of 50 °C.•PtGR folds with a typical crystal structure of d-2-hydroxy-acid dehydrogenase.•A cationic cluster help PtGR to prefer NADPH rather than NADH as coenzyme.

A glyoxylate reductase gene (PtGR) from the fungus Paecilomyces thermophila was cloned and expressed in Escherichia coli. PtGR was biochemically and structurally characterized. PtGR has an open reading frame of 993 bp encoding 330 amino acids. The deduced amino acid sequence has low similarities to the reported glyoxylate reductases. The purified PtGR forms a homodimer. PtGR displayed an optimum pH of 7.5 and broad pH stability (pH 4.5–10). It exhibited an optimal temperature of 50 °C and was stable up to 50 °C. PtGR was found to be highly specific for glyoxylate, but it showed no detectable activity with 4-methyl-2-oxopentanoate, phenylglyoxylate, pyruvate, oxaloacetate and α-ketoglutarate. PtGR prefered NADPH rather than NADH as an electron donor. Moreover, the crystal structure of PtGR was determined at 1.75 Å resolution. The overall structure of apo-PtGR monomer adopts the typical d-2-hydroxy-acid dehydrogenase fold with a “closed” conformation unexpectedly. The coenzyme specificity is provided by a cationic cluster consisting of N184, R185, and N186 structurally. These structural observations could explain its different coenzyme and substrate specificity.

Keywords
AtGLYR1, glyoxylate reducatase from Arabidopsis thaliana; CbBH(P)PR, hydroxyl(phenyl)pyruvate reductase from Coleus blumei Benth; hGRHPR, human glyoxylate reductase/hydroxypyruvate reductase; IDA, iminodiacetic acid; IPTG, isopropyl β-d-thiogalactopyranos
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Characterization and crystal structure of a first fungal glyoxylate reductase from Paecilomyes thermophila
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 60, 10 June 2014, Pages 72–79
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
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Price after discount Only $4.95
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Any Questions? feel free to contact us