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Title: Cofactor engineering of ketol-acid reductoisomerase (IlvC) and alcohol dehydrogenase (YqhD) improves the fusel alcohol yield in algal protein anaerobic fermentation

Abstract

Recently the feasibility of conversion of algal protein to mixed alcohols has been demonstrated with an engineered E.coli strain, enabling comprehensive utilization of the biomass for biofuel applications. However, the yield and titers of mixed alcohol production must be improved for market adoption. A major limiting factor for achieving the necessary yield and titer improvements is cofactor imbalance during the fermentation of algal protein. To resolve this problem, a directed evolution approach was applied to modify the cofactor specificity of two key enzymes (IlvC and YqhD) from NADPH to NADH in the mixed alcohol metabolic pathway. Using high throughput screening, more than 20 YqhD mutants were identified to show activity on NADH as a cofactor. Of these 20 mutants, the top five of YqhD mutants were selected for combination with two IlvC mutants with NADH as a cofactor for the modification of the protein conversion strain. The combination of the IlvC and YqhD mutants yielded a refined E.coli strain, subtype AY3, with increased fusel alcohol yield of ~60% compared to wild type under anaerobic fermentation on amino acid mixtures. When applied to real algal protein hydrolysates, the strain AY3 produced 100% and 38% more total mixed alcohols than the wildmore » type strain on two different algal hydrolysates, respectively. The results indicate that cofactor engineering is a promising approach to improve the feasibility of bioconversion of algal protein into mixed alcohols as advanced biofuels.« less

Authors:
 [1];  [1];  [1];  [1]
  1. Sandia National Lab. (SNL-CA), Livermore, CA (United States)
Publication Date:
Research Org.:
Sandia National Lab. (SNL-CA), Livermore, CA (United States)
Sponsoring Org.:
USDOE Office of Energy Efficiency and Renewable Energy (EERE)
OSTI Identifier:
1338336
Alternate Identifier(s):
OSTI ID: 1402509
Report Number(s):
SAND2016-2002J
Journal ID: ISSN 2211-9264; PII: S2211926416302715
Grant/Contract Number:  
AC04-94AL85000; ACO4-94AL85000; 26336
Resource Type:
Accepted Manuscript
Journal Name:
Algal Research
Additional Journal Information:
Journal Volume: 19; Journal Issue: C; Journal ID: ISSN 2211-9264
Publisher:
Elsevier
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; directed evolution; isobutanol, 2/3-methyl butanol; algae protein bioconversion; cofactor engineering; algal biofuel

Citation Formats

Wu, Weihua, Tran-Gyamfi, Mary Bao, Jaryenneh, James Dekontee, and Davis, Ryan W. Cofactor engineering of ketol-acid reductoisomerase (IlvC) and alcohol dehydrogenase (YqhD) improves the fusel alcohol yield in algal protein anaerobic fermentation. United States: N. p., 2016. Web. https://doi.org/10.1016/j.algal.2016.08.013.
Wu, Weihua, Tran-Gyamfi, Mary Bao, Jaryenneh, James Dekontee, & Davis, Ryan W. Cofactor engineering of ketol-acid reductoisomerase (IlvC) and alcohol dehydrogenase (YqhD) improves the fusel alcohol yield in algal protein anaerobic fermentation. United States. https://doi.org/10.1016/j.algal.2016.08.013
Wu, Weihua, Tran-Gyamfi, Mary Bao, Jaryenneh, James Dekontee, and Davis, Ryan W. Wed . "Cofactor engineering of ketol-acid reductoisomerase (IlvC) and alcohol dehydrogenase (YqhD) improves the fusel alcohol yield in algal protein anaerobic fermentation". United States. https://doi.org/10.1016/j.algal.2016.08.013. https://www.osti.gov/servlets/purl/1338336.
@article{osti_1338336,
title = {Cofactor engineering of ketol-acid reductoisomerase (IlvC) and alcohol dehydrogenase (YqhD) improves the fusel alcohol yield in algal protein anaerobic fermentation},
author = {Wu, Weihua and Tran-Gyamfi, Mary Bao and Jaryenneh, James Dekontee and Davis, Ryan W.},
abstractNote = {Recently the feasibility of conversion of algal protein to mixed alcohols has been demonstrated with an engineered E.coli strain, enabling comprehensive utilization of the biomass for biofuel applications. However, the yield and titers of mixed alcohol production must be improved for market adoption. A major limiting factor for achieving the necessary yield and titer improvements is cofactor imbalance during the fermentation of algal protein. To resolve this problem, a directed evolution approach was applied to modify the cofactor specificity of two key enzymes (IlvC and YqhD) from NADPH to NADH in the mixed alcohol metabolic pathway. Using high throughput screening, more than 20 YqhD mutants were identified to show activity on NADH as a cofactor. Of these 20 mutants, the top five of YqhD mutants were selected for combination with two IlvC mutants with NADH as a cofactor for the modification of the protein conversion strain. The combination of the IlvC and YqhD mutants yielded a refined E.coli strain, subtype AY3, with increased fusel alcohol yield of ~60% compared to wild type under anaerobic fermentation on amino acid mixtures. When applied to real algal protein hydrolysates, the strain AY3 produced 100% and 38% more total mixed alcohols than the wild type strain on two different algal hydrolysates, respectively. The results indicate that cofactor engineering is a promising approach to improve the feasibility of bioconversion of algal protein into mixed alcohols as advanced biofuels.},
doi = {10.1016/j.algal.2016.08.013},
journal = {Algal Research},
number = C,
volume = 19,
place = {United States},
year = {2016},
month = {8}
}

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Cited by: 4 works
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    Works referencing / citing this record:

    Bioconversion of distillers’ grains hydrolysates to advanced biofuels by an Escherichia coli co-culture
    journal, November 2017


    Characterization of four endophytic fungi as potential consolidated bioprocessing hosts for conversion of lignocellulose into advanced biofuels
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    • Wu, Weihua; Davis, Ryan W.; Tran-Gyamfi, Mary Bao
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    Development of algae biorefinery concepts for biofuels and bioproducts; a perspective on process-compatible products and their impact on cost-reduction
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