Crystal Structure and Biophysical Analysis of Furfural-Detoxifying Aldehyde Reductase from Clostridium beijerinckii

Many aldehydes, such as furfural, are present in high quantities in lignocellulose lysates and are fermentation inhibitors, which makes biofuel production from this abundant carbon source extremely challenging. Cbei_3974 has recently been identified as an aldo-keto reductase responsible for partial...

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Veröffentlicht in:Applied and environmental microbiology 2019-08, Vol.85 (15), p.1
Hauptverfasser: Scott, Alan F, Cresser-Brown, Joel, Williams, Thomas L, Rizkallah, Pierre J, Jin, Yi, Luk, Louis Y-P, Allemann, Rudolf K
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container_issue 15
container_start_page 1
container_title Applied and environmental microbiology
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creator Scott, Alan F
Cresser-Brown, Joel
Williams, Thomas L
Rizkallah, Pierre J
Jin, Yi
Luk, Louis Y-P
Allemann, Rudolf K
description Many aldehydes, such as furfural, are present in high quantities in lignocellulose lysates and are fermentation inhibitors, which makes biofuel production from this abundant carbon source extremely challenging. Cbei_3974 has recently been identified as an aldo-keto reductase responsible for partial furfural resistance in Rational engineering of this enzyme could enhance the furfural tolerance of this organism, thereby improving biofuel yields. We report an extensive characterization of Cbei_3974 and a single-crystal X-ray structure of Cbei_3974 in complex with NADPH at a resolution of 1.75 Å. Docking studies identified residues involved in substrate binding, and an activity screen revealed the substrate tolerance of the enzyme. Hydride transfer, which is partially rate limiting under physiological conditions, occurs from the pro- hydrogen of NADPH. Enzyme isotope labeling revealed a temperature-independent enzyme isotope effect of unity, indicating that the enzyme does not use dynamic coupling for catalysis and suggesting that the active site of the enzyme is optimally configured for catalysis with the substrate tested. Here we report the crystal structure and biophysical properties of an aldehyde reductase that can detoxify furfural, a common inhibitor of biofuel fermentation found in lignocellulose lysates. The data contained here will serve as a guide for protein engineers to develop improved enzyme variants that would impart furfural resistance to the microorganisms used in biofuel production and thus lead to enhanced biofuel yields from this sustainable resource.
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Cbei_3974 has recently been identified as an aldo-keto reductase responsible for partial furfural resistance in Rational engineering of this enzyme could enhance the furfural tolerance of this organism, thereby improving biofuel yields. We report an extensive characterization of Cbei_3974 and a single-crystal X-ray structure of Cbei_3974 in complex with NADPH at a resolution of 1.75 Å. Docking studies identified residues involved in substrate binding, and an activity screen revealed the substrate tolerance of the enzyme. Hydride transfer, which is partially rate limiting under physiological conditions, occurs from the pro- hydrogen of NADPH. Enzyme isotope labeling revealed a temperature-independent enzyme isotope effect of unity, indicating that the enzyme does not use dynamic coupling for catalysis and suggesting that the active site of the enzyme is optimally configured for catalysis with the substrate tested. Here we report the crystal structure and biophysical properties of an aldehyde reductase that can detoxify furfural, a common inhibitor of biofuel fermentation found in lignocellulose lysates. 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Here we report the crystal structure and biophysical properties of an aldehyde reductase that can detoxify furfural, a common inhibitor of biofuel fermentation found in lignocellulose lysates. The data contained here will serve as a guide for protein engineers to develop improved enzyme variants that would impart furfural resistance to the microorganisms used in biofuel production and thus lead to enhanced biofuel yields from this sustainable resource.</abstract><cop>United States</cop><pub>American Society for Microbiology</pub><pmid>31101612</pmid><doi>10.1128/AEM.00978-19</doi><orcidid>https://orcid.org/0000-0002-5008-8958</orcidid><oa>free_for_read</oa></addata></record>
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subjects Aldehyde reductase
Aldehydes
Aldo-keto reductase
Bacteria
Biodiesel fuels
Biofuels
Biomass
Carbon sources
Catalysis
Clostridium beijerinckii
Crystal structure
Docking
Enzymes
Enzymology and Protein Engineering
Fermentation
Fuels
Furfural
Isotope effect
Lignocellulose
Lysates
NADP
Reductases
Single crystals
Substrates
title Crystal Structure and Biophysical Analysis of Furfural-Detoxifying Aldehyde Reductase from Clostridium beijerinckii
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