ADH from Rhodococcus ruber expressed in Arxula adeninivorans for the synthesis of 1-(S)-phenylethanol

Rauter, Marion; Kasprzak, Jakub; Becker, K.; Baronian, Keith; Bode, R.; Kunze, Gotthard; Vorbrodt, H.-Matthias

doi:10.1016/j.molcatb.2014.02.015

Cited by 15 publications

(10 citation statements)

References 25 publications

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“…T h e A D H a c t i v i t i e s a t t a i n e d b y t r a n s g e n i c A. adeninivorans strains were 2 to 4 times higher than wildtype L. kefir (1200 U l −1 ADH) (Hummel 1997) and 15 to 28 times higher than RrADH producing A. adeninivorans strain (183 U l −1 ) (Rauter et al 2014a). Regenerating enzyme activity was 2.7 to 1.3 times lower with 170 or 370 U l −1 , respectively (Rauter et al 2014a).…”

Section: Discussionmentioning

confidence: 89%

“…Regenerating enzyme activity was 2.7 to 1.3 times lower with 170 or 370 U l −1 , respectively (Rauter et al 2014a). Functionality of the different enzymes was not only determined by observing NADH oxidation assays, but also by product analysis with GC-FID.…”

Section: Discussionmentioning

confidence: 99%

“…Xplor2.2-TEF1-(2)LbADH-PHO5 and Xplor2.4-TEF1-BpG6PDH-PHO5 or Xplor2.4-TEF1-BmGDH-PHO5 (Rauter et al 2014a) were used to coexpress LbADH and BmGDH (accession no. LK055286) or BpG6PDH gene in A. adeninivorans MS1006.…”

Section: Construction Of Lbadh and Bpg6pdh Expression Plasmidsmentioning

confidence: 99%

“…BpG6PDH and LbADH producing yeast strains were produced as described by Rauter et al (2014a). This process involves the ligation of the LbADH gene and ATRP1m or BpG6PDH gene and AADE2 selection marker module.…”

Section: Expression Of Lbadh and Bpg6pdh In A Adeninivoransmentioning

confidence: 99%

“…adeninivorans has already been used for the expression of the ADH gene of Rhodococcus ruber and a recombinant strain coexpressing RrADH and Bacillus megaterium GDH was used for the synthesis of enantiomerically pure 1-(S)-phenylethanol (Giersberg et al 2012;Rauter et al 2014a). …”

Section: Introductionmentioning

confidence: 99%

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Coexpression of Lactobacillus brevis ADH with GDH or G6PDH in Arxula adeninivorans for the synthesis of 1-(R)-phenylethanol

Rauter¹,

Prokoph

Kasprzak

et al. 2014

Appl Microbiol Biotechnol

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The yeast Arxula adeninivorans was used for the overexpression of an ADH gene of Lactobacillus brevis coding for (R)-specific alcohol dehydrogenase (LbADH) to synthesise enantiomerically pure 1-(R)-phenylethanol. Glucose dehydrogenase gene from Bacillus megaterium (BmGDH) or glucose 6-phosphate dehydrogenase of Bacillus pumilus (BpG6PDH) were coexpressed in Arxula to regenerate the cofactor NADPH by oxidising glucose or glucose 6-phosphate. The yeast strain expressing LbADH and BpG6PDH produced 5200 U l(-1) ADH and 370 U l(-1) G6PDH activity, whereas the strain expressing LbADH and BmGDH produced 2700 U l(-1) ADH and 170 U l(-1) GDH activity. However, the crude extract of both strains reduced 40 mM acetophenone to pure 1-(R)-phenylethanol with an enantiomeric excess (ee) of >99 % in 60 min without detectable by-products. An increase in yield was achieved using immobilised crude extracts (IEs), Triton X-100 permeabilised cells (PCs) and permeabilised immobilised cells (PICs) with PICs being most stable with GDH regeneration over 52 cycles. Even though the activity and synthesis rate of 1-(R)-phenylethanol with the BpG6PDH and LbADH coexpressing strain was higher, the BmGDH-LbADH strain was more stable over successive reaction cycles. This, combined with its higher total turnover number (TTN) of 391 mol product per mole NADP(+), makes it the preferred strain for continuous reaction systems. The initial non-optimised semi-continuous reaction produced 9.74 g l(-1) day(-1) or 406 g kg(-1) dry cell weight (dcw) day(-1) isolated 1-(R)-phenylethanol with an ee of 100 % and a TTN of 206 mol product per mole NADP(+). In conclusion, A. adeninivorans is a promising host for LbADH and BpG6PDH or BmGDH production and offers a simple method for the production of enantiomerically pure alcohols.

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Section: Discussionmentioning

confidence: 89%

Section: Discussionmentioning

confidence: 99%

Section: Construction Of Lbadh and Bpg6pdh Expression Plasmidsmentioning

confidence: 99%

Section: Expression Of Lbadh and Bpg6pdh In A Adeninivoransmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Coexpression of Lactobacillus brevis ADH with GDH or G6PDH in Arxula adeninivorans for the synthesis of 1-(R)-phenylethanol

Rauter¹,

Prokoph

Kasprzak

et al. 2014

Appl Microbiol Biotechnol

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Blastobotrys (Arxula) adeninivorans: a promising alternative yeast for biotechnology and basic research

Malak

Baronian

Kunze

2016

Yeast

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Blastobotrys adeninivorans (syn. Arxula adeninivorans) is a non-conventional, non-pathogenic, imperfect, haploid yeast, belonging to the subphylum Saccharomycotina, which has to date received comparatively little attention from researchers. It possesses unusual properties such as thermo- and osmotolerance, and a broad substrate spectrum. Depending on the cultivation temperature B. (A.) adeninivorans exhibits different morphological forms and various post-translational modifications and protein expression properties that are strongly correlated with the morphology. The genome has been completely sequenced and, in addition, there is a well-developed transformation/expression platform, which makes rapid, simple gene manipulations possible. This yeast species is a very good host for homologous and heterologous gene expression and is also a useful gene donor. Blastobotrys (A.) adeninivorans is able to use a very wide range of substrates as carbon and/or nitrogen sources and is an interesting organism owing to the presence of many metabolic pathways, for example degradation of n-butanol, purines and tannin. In addition, its unusual properties and robustness make it a useful bio-component for whole cell biosensors. There are currently a number of products on the market produced by B. (A.) adeninivorans and further investigation may contribute further innovative solutions for current challenges that exist in the biotechnology industry. Additionally it may become a useful alternative to existing commercial yeast strains and as a model organism in research. In this review we present information relevant to the exploitation of B. (A.) adeninivorans in research and industrial settings. Copyright © 2016 John Wiley & Sons, Ltd.

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Selection of the Optimal Yeast Host for the Synthesis of Recombinant Enzymes

Bischoff

Giersberg

Matthes

et al. 2019

Methods in Molecular Biology

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ADH from Rhodococcus ruber expressed in Arxula adeninivorans for the synthesis of 1-(S)-phenylethanol

Cited by 15 publications

References 25 publications

Coexpression of Lactobacillus brevis ADH with GDH or G6PDH in Arxula adeninivorans for the synthesis of 1-(R)-phenylethanol

Coexpression of Lactobacillus brevis ADH with GDH or G6PDH in Arxula adeninivorans for the synthesis of 1-(R)-phenylethanol

Blastobotrys (Arxula) adeninivorans: a promising alternative yeast for biotechnology and basic research

Selection of the Optimal Yeast Host for the Synthesis of Recombinant Enzymes

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