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Characterization of alcohol dehydrogenase 1 and 3 from Neurospora crassa FGSC2489

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Abstract

Alcohol dehydrogenase (ADH) is a key enzyme in the production and utilization of alcohols. Some also catalyze the formation of carboxylate esters from alcohols and aldehydes. The ADH1 and ADH3 genes of Neurospora crassa FGSC2489 were cloned and expressed in recombinant Escherichia coli to investigate their alcohol dehydrogenation and carboxylate ester formation abilities. Homology analysis and sequence alignment of amino acid sequence indicated that ADH1 and ADH3 of N. crassa contained a zinc-binding consensus sequence and a NAD+-binding motif and showed 54–75% identity with fungi ADHs. N. crassa ADH1 was expressed in E. coli to give a specific activity of 289 ± 9 mU/mg using ethanol and NAD+ as substrate and cofactor, respectively. Corresponding experiments on the expression and activity of ADH3 gave 4 mU/mg of specific activity. N. crassaADH1 preferred primary alcohols containing C3–C8 carbons to secondary alcohols such as 2-propanol and 2-butanol. N. crassaADH1 possessed 5.3 mU/mg of specific carboxylate ester-forming activity accumulating 0.4 mM of ethyl acetate in 18 h. Substrate specificity of various linear alcohols and aldehydes indicated that short chain-length alcohols and aldehydes were good substrates for carboxylate ester production. N. crassaADH1 was a primary alcohol dehydrogenase using cofactor NAD+ preferably and possessed carboxylate ester-forming activity with short chain alcohols and aldehydes.

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Acknowledgment

This study was supported by the Korea Research Foundation Grant (KRF-2004-000-10255-0) and USDA-NSF Interagency Metabolic Engineering Program: USDA 2002-35505-11638 and NSF BES 0118815.

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  1. Yong-Cheol Park

    Present address: Center for Agricultural Biomaterials, Seoul National University, Seoul, 151-921, South Korea

Authors and Affiliations

  1. Department of Biochemistry and Cell Biology, Rice University, 6100 Main St., Houston, TX, 77005, USA

    Yong-Cheol Park & George N. Bennett

  2. Department of Chemical Engineering, Rice University, 6100 Main St., Houston, TX, 77005, USA

    Ka-Yiu San

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  1. Yong-Cheol Park
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  2. Ka-Yiu San
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  3. George N. Bennett
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Correspondence to George N. Bennett.

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Park, YC., San, KY. & Bennett, G.N. Characterization of alcohol dehydrogenase 1 and 3 from Neurospora crassa FGSC2489. Appl Microbiol Biotechnol 76, 349–356 (2007). https://doi.org/10.1007/s00253-007-0998-5

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  • DOI: https://doi.org/10.1007/s00253-007-0998-5

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