Engineering of xylose reductase and overexpression of xylitol dehydrogenase and xylulokinase improves xylose alcoholic fermentation in the thermotolerant yeast Hansenula polymorpha

Dmytruk, Olena V.; Dmytruk, Kostyantyn V.; Abbas, Charles A.; Voronovsky, Andriy Y.; Sibirny, Andriy А.

doi:10.1186/1475-2859-7-21

Cited by 47 publications

(24 citation statements)

References 33 publications

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“…Several different strategies have been developed to produce ethanol from xylose with genetically engineered microorganisms, using either thermotolerant yeast and thermophilic bacteria in simultaneous saccharification and fermentation processes [9,36] or mesophilic bacteria with high growth rates [26].…”

Section: Introductionmentioning

confidence: 99%

Effect of controlled oxygen limitation on Candida shehatae physiology for ethanol production from xylose and glucose

Fromanger

Guillouet

Uribelarrea

et al. 2010

J Ind Microbiol Biotechnol

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Carbon distribution and kinetics of Candida shehatae were studied in fed-batch fermentation with xylose or glucose (separately) as the carbon source in mineral medium. The fermentations were carried out in two phases, an aerobic phase dedicated to growth followed by an oxygen limitation phase dedicated to ethanol production. Oxygen limitation was quantified with an average specific oxygen uptake rate (OUR) varying between 0.30 and 2.48 mmolO(2) g dry cell weight (DCW)(-1) h(-1), the maximum value before the aerobic shift. The relations among respiration, growth, ethanol production and polyol production were investigated. It appeared that ethanol was produced to provide energy, and polyols (arabitol, ribitol, glycerol and xylitol) were produced to reoxidize NADH from assimilatory reactions and from the co-factor imbalance of the two-first enzymatic steps of xylose uptake. Hence, to manage carbon flux to ethanol production, oxygen limitation was a major controlled parameter; an oxygen limitation corresponding to an average specific OUR of 1.19 mmolO(2) g DCW(-1) h(-1) allowed maximization of the ethanol yield over xylose (0.327 g g(-1)), the average productivity (2.2 g l(-1) h(-1)) and the ethanol final titer (48.81 g l(-1)). For glucose fermentation, the ethanol yield over glucose was the highest (0.411 g g(-1)) when the specific OUR was low, corresponding to an average specific OUR of 0.30 mmolO(2) g DCW(-1) h(-1), whereas the average ethanol productivity and ethanol final titer reached the maximum values of 1.81 g l(-1) h(-1) and 54.19 g l(-1) when the specific OUR was the highest.

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Section: Introductionmentioning

confidence: 99%

Effect of controlled oxygen limitation on Candida shehatae physiology for ethanol production from xylose and glucose

Fromanger

Guillouet

Uribelarrea

et al. 2010

J Ind Microbiol Biotechnol

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“…Though maximal ethanol yield from xylose achieved by constructed strains was low (0.9 g of ethanol per liter), this number can be significantly increased using approaches of metabolic engineering of H. polymorpha recently developed in our laboratory. Among the approaches successfully used and reported from our lab to improve ethanol fermentation from xylose at high temperature are the: (i) overexpression of bacterial xylose isomerase and own xylulokinase (Dmytruk et al, 2008a); (ii) overexpression of engineered xylose reductase, xylitol dehydrogenase, and xylulokinase (Dmytruk et al, 2008b); (iii) overexpression of pyruvate decarboxylase (Ishchuk et al, 2008). We suggest that application of these approaches to the strains constructed in the current work, will further improve xylose alcoholic fermentation at 508C, which will result in development of the robust strains suitable for the process of SSF.…”

Section: Discussionmentioning

confidence: 99%

Construction of Hansenula polymorpha strains with improved thermotolerance

Ishchuk

Voronovsky

Abbas

et al. 2009

Biotech & Bioengineering

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The methylotrophic yeast Hansenula polymorpha has the potential to be used in the process of simultaneous saccharification and fermentation (SSF) of xylan derived xylose at elevated temperatures. To improve parameters of high-temperature resistance and high-temperature fermentation of H. polymorpha, strains carrying deletion of acid trehalase gene (ATH1) and overexpressing genes coding for heat-shock proteins Hsp16p and Hsp104p were constructed. Results indicate that the corresponding recombinant strains have up to 12-fold increased tolerance to heat-shock treatment. The deletion of ATH1 gene and constitutive expression of HSP16 and HSP104 resulted in up to 5.8-fold improvement of ethanol production from xylose at 50 degrees C. Although the maximum ethanol concentration achieved from xylose was 0.9 g L(-1), our model H. polymorpha strains with elevated thermotolerance can be further modified by metabolic engineering to construct improved high-temperature ethanol producers from this pentose.

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“…Moreover, peculiar physiological characteristics of H. polymorpha , such as resistance to heavy metals, oxidative stress and heat, make this yeast attractive for several biotechnological purposes (Blazhenko et al , 2006). Recently, the capability of this thermotolerant yeast in alcoholic fermentation of xylose at elevated temperatures has drawn attention to the yeast as a good candidate for the development of an efficient process for simultaneous saccharification and fermentation (Dmytruk et al , 2008). With the completion of H. polymorpha genome sequencing (Ramezani‐Rad et al , 2003), post‐genomic approaches such as transcriptome (Park et al , 2007) and proteome analysis (Kim et al , 2004b) are now feasible and enable more systematic strategies for metabolic engineering and process improvement.…”

Section: Introductionmentioning

confidence: 99%

New selectable host–marker systems for multiple genetic manipulations based on TRP1, MET2 and ADE2 in the methylotrophic yeast Hansenula polymorpha

Cheon

Choo

Ubiyvovk

et al. 2009

Yeast

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Interest has been increasing in the thermotolerant methylotrophic yeast Hansenula polymorpha as a useful system for fundamental research and applied purposes. Only a few genetic marker genes and auxotrophic hosts are yet available for this yeast. Here we isolated and developed H. polymorpha TRP1, MET2 and ADE2 genes as selectable markers for multiple genetic manipulations. The H. polymorpha TRP1 (HpTRP1 ), MET2 (HpMET2 ) and ADE2 (HpADE2 ) genes were sequentially disrupted, using an HpURA3 pop-out cassette in H. polymorpha to generate a series of new multiple auxotrophic strains, including up to a quintuple auxotrophic strain. Unexpectedly, the HpTRP1 deletion mutants required additional tryptophan supplementation for their full growth, even on complex media such as YPD. Despite the clearly increased resistance to 5-fluoroanthranilic acid of the HpTRP1 deletion mutants, the HpTRP1 blaster cassette does not appear to be usable as a counter-selection marker in H. polymorpha. Expression vectors carrying HpADE2, HpTRP1 or HpMET2 with their own promoters and terminators as selectable markers were constructed and used to co-transform the quintuple auxotrophic strain for the targeted expression of a heterologous gene, Aspergillus saitoi MsdS, at the ER, the Golgi and the cell surface, respectively. The nucleotide sequences presented here were submitted to GenBank under Accession Nos AY795576 (HpTRP1 ), FJ226453 (HpMET2 ) and FJ493241 (HpADE2 ), respectively.

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Engineering of xylose reductase and overexpression of xylitol dehydrogenase and xylulokinase improves xylose alcoholic fermentation in the thermotolerant yeast Hansenula polymorpha

Cited by 47 publications

References 33 publications

Effect of controlled oxygen limitation on Candida shehatae physiology for ethanol production from xylose and glucose

Effect of controlled oxygen limitation on Candida shehatae physiology for ethanol production from xylose and glucose

Construction of Hansenula polymorpha strains with improved thermotolerance

New selectable host–marker systems for multiple genetic manipulations based on TRP1, MET2 and ADE2 in the methylotrophic yeast Hansenula polymorpha

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