Large-scale production of tannase using the yeast Arxula adeninivorans

Böer, Erik; Breuer, Friederike Sophie; Weniger, Michael; Denter, Sylvia; Piontek, Michael; Kunze, Gotthard

doi:10.1007/s00253-011-3320-5

Cited by 30 publications

(14 citation statements)

References 43 publications

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“…Different secretory enzymes have already been reported to be produced by A. adeninivorans, showing the great biotechnological potential of this yeast species (26,40). However, the secretion of cutinases or cutinase-like enzymes by A. adeninivorans has not been reported to date.…”

Section: Discussionmentioning

confidence: 95%

Three New Cutinases from the Yeast Arxula adeninivorans That Are Suitable for Biotechnological Applications

Bischoff

Litwińska

Cordes³

et al. 2015

Appl Environ Microbiol

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The genes ACUT1, ACUT2, and ACUT3, encoding cutinases, were selected from the genomic DNA of Arxula adeninivorans LS3. The alignment of the amino acid sequences of these cutinases with those of other cutinases or cutinase-like enzymes from different fungi showed that they all had a catalytic S-D-H triad with a conserved G-Y-S-Q-G domain. All three genes were overexpressed in A. adeninivorans using the strong constitutive TEF1 promoter. Recombinant 6؋ His (6h)-tagged cutinase 1 protein (p) from A. adeninivorans LS3 (Acut1-6hp), Acut2-6hp, and Acut3-6hp were produced and purified by immobilized-metal ion affinity chromatography and biochemically characterized using p-nitrophenyl butyrate as the substrate for standard activity tests. All three enzymes from A. adeninivorans were active from pH 4.5 to 6.5 and from 20 to 30°C. They were shown to be unstable under optimal reaction conditions but could be stabilized using organic solvents, such as polyethylene glycol 200 (PEG 200), isopropanol, ethanol, or acetone. PEG 200 (50%, vol/vol) was found to be the best stabilizing agent for all of the cutinases, and acetone greatly increased the half-life and enzyme activity (up to 300% for Acut3-6hp). The substrate spectra for Acut1-6hp, Acut2-6hp, and Acut3-6hp were quite similar, with the highest activity being for short-chain fatty acid esters of p-nitrophenol and glycerol. Additionally, they were found to have polycaprolactone degradation activity and cutinolytic activity against cutin from apple peel. The activity was compared with that of the 6؋ His-tagged cutinase from Fusarium solani f. sp. pisi (FsCut-6hp), also expressed in A. adeninivorans, as a positive control. A fed-batch cultivation of the best Acut2-6hp-producing strain, A. adeninivorans G1212/YRC102-ACUT2-6H, was performed and showed that very high activities of 1,064 U ml ؊1 could be achieved even with a nonoptimized cultivation procedure.

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Section: Discussionmentioning

confidence: 95%

Three New Cutinases from the Yeast Arxula adeninivorans That Are Suitable for Biotechnological Applications

Bischoff

Litwińska

Cordes³

et al. 2015

Appl Environ Microbiol

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“…First, transgenic tannase producer strains were constructed with a constitutively expressed ATAN1 module integrated into a chromosome. In fed-batch fermentation experiments, the transgenic strain produced 51,900 U/L of tannase activity after 42 h with a dry cell weight of 162 g/L [1]. …”

Section: Resultsmentioning

confidence: 99%

“…This species is currently exploited as biocatalyst for the synthesis of various biotechnological products such as tannases [1], 1-(S)-phenylethanol [2] or β-D-galactopyranoside [3], for the production of food with low purine content [4], and for the detection of estrogenic activity in various aqueous media [5,6]. It is also used as a host for the production of recombinant proteins, and as a donor for genes encoding valuable products [7,8].…”

Section: Introductionmentioning

confidence: 99%

The complete genome of Blastobotrys (Arxula) adeninivorans LS3 - a yeast of biotechnological interest

Kunze

Gaillardin

Czernicka³

et al. 2014

Biotechnol Biofuels

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BackgroundThe industrially important yeast Blastobotrys (Arxula) adeninivorans is an asexual hemiascomycete phylogenetically very distant from Saccharomyces cerevisiae. Its unusual metabolic flexibility allows it to use a wide range of carbon and nitrogen sources, while being thermotolerant, xerotolerant and osmotolerant.ResultsThe sequencing of strain LS3 revealed that the nuclear genome of A. adeninivorans is 11.8 Mb long and consists of four chromosomes with regional centromeres. Its closest sequenced relative is Yarrowia lipolytica, although mean conservation of orthologs is low. With 914 introns within 6116 genes, A. adeninivorans is one of the most intron-rich hemiascomycetes sequenced to date. Several large species-specific families appear to result from multiple rounds of segmental duplications of tandem gene arrays, a novel mechanism not yet described in yeasts. An analysis of the genome and its transcriptome revealed enzymes with biotechnological potential, such as two extracellular tannases (Atan1p and Atan2p) of the tannic-acid catabolic route, and a new pathway for the assimilation of n-butanol via butyric aldehyde and butyric acid.ConclusionsThe high-quality genome of this species that diverged early in Saccharomycotina will allow further fundamental studies on comparative genomics, evolution and phylogenetics. Protein components of different pathways for carbon and nitrogen source utilization were identified, which so far has remained unexplored in yeast, offering clues for further biotechnological developments. In the course of identifying alternative microorganisms for biotechnological interest, A. adeninivorans has already proved its strengthened competitiveness as a promising cell factory for many more applications.

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“…Among the few recombinant enzymes that have been expressed in A. adeninivorans, a tannase has been successfully over-produced using various plasmids carrying one or two expression modules. 157 S. pombe is an attractive host for the expression of recombinant enzymes. An endopolygalacturonase from S. cerevisiae PGU1 was successfully expressed in S. pombe.…”

Section: Candida Boidiniimentioning

confidence: 99%

How to achieve high-level expression of microbial enzymes

et al. 2013

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Microbial enzymes have been used in a large number of fields, such as chemical, agricultural and biopharmaceutical industries. The enzyme production rate and yield are the main factors to consider when choosing the appropriate expression system for the production of recombinant proteins. Recombinant enzymes have been expressed in bacteria (e.g., Escherichia coli, Bacillus and lactic acid bacteria), filamentous fungi (e.g., Aspergillus) and yeasts (e.g., Pichia pastoris). The favorable and very advantageous characteristics of these species have resulted in an increasing number of biotechnological applications. Bacterial hosts (e.g., E. coli) can be used to quickly and easily overexpress recombinant enzymes; however, bacterial systems cannot express very large proteins and proteins that require post-translational modifications. The main bacterial expression hosts, with the exception of lactic acid bacteria and filamentous fungi, can produce several toxins which are not compatible with the expression of recombinant enzymes in food and drugs. However, due to the multiplicity of the physiological impacts arising from high-level expression of genes encoding the enzymes and expression hosts, the goal of overproduction can hardly be achieved, and therefore, the yield of recombinant enzymes is limited. In this review, the recent strategies used for the high-level expression of microbial enzymes in the hosts mentioned above are summarized and the prospects are also discussed. We hope this review will contribute to the development of the enzyme-related research field.

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Large-scale production of tannase using the yeast Arxula adeninivorans

Cited by 30 publications

References 43 publications

Three New Cutinases from the Yeast Arxula adeninivorans That Are Suitable for Biotechnological Applications

Three New Cutinases from the Yeast Arxula adeninivorans That Are Suitable for Biotechnological Applications

The complete genome of Blastobotrys (Arxula) adeninivorans LS3 - a yeast of biotechnological interest

How to achieve high-level expression of microbial enzymes

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