Publications

• Alternaria alternata NADP-dependent mannitol dehydrogenase is an important fungal allergen.

  Schneider P.B., Denk U., Breitenbach M., Richter K., Schmid-Grendelmeier P., Nobbe S., Himly M., Mari A., Ebner C., Simon-Nobbe B.

  <h4>Background</h4>Alternaria alternata is one of the most important allergenic fungi worldwide. Mannitol dehydrogenase (MtDH) has previously been shown to be a major allergen of Cladosporium herbarum and cross-reactivity has been demonstrated for several fungal allergens.<h4>Objective</h4>The pre ... >> More

  <h4>Background</h4>Alternaria alternata is one of the most important allergenic fungi worldwide. Mannitol dehydrogenase (MtDH) has previously been shown to be a major allergen of Cladosporium herbarum and cross-reactivity has been demonstrated for several fungal allergens.<h4>Objective</h4>The present study's objective was to clone the MtDH from an A. alternata cDNA library, express and purify the recombinant non-fusion protein and test its IgE-binding properties. Methods A cDNA library prepared from A. alternata hyphae and spores was screened for mannitol dehydrogenase by DNA hybridization with the radioactively labelled C. herbarum homologue as a probe. The resulting clone was sequenced and heterologously expressed in Escherichia coli as a recombinant non-fusion protein, which was purified to homogeneity and analysed for its IgE-binding capacity.<h4>Results</h4>The coding sequence of the full-length cDNA clone comprises 798 bp encoding a protein with a molecular mass of 28.6 kDa and a predicted pI of 5.88. Protein sequence analysis revealed an identity of 75% and a homology of 86% between the MtDHs of A. alternata and C. herbarum. The functional mannitol dehydrogenase was expressed in the E. coli strain BL21(DE3) transformed with the vector pMW172 and purified to homogeneity. The enzyme catalyses the NADPH-dependent conversion of d-fructose to d-mannitol. In IgE-ELISA and immunoblots, MtDH is recognized by 41% of A. alternata-allergic patients. In vivo immunoreactivity of the recombinant MtDH was verified by skin prick testing. Finally, inhibition-ELISA experiments confirmed cross-reactivity between the MtDHs of A. alternata and C. herbarum.<h4>Conclusion</h4>Mannitol dehydrogenase (Alt a 8) represents an important new allergen of the ascomycete A. alternata that might be suitable for improving diagnostic and therapeutic procedures. << Less

  Clin. Exp. Allergy 36:1513-1524(2006) [PubMed] [EuropePMC]

• The Hypocrea jecorina (syn. Trichoderma reesei) lxr1 gene encodes a D-mannitol dehydrogenase and is not involved in L-arabinose catabolism.

  Metz B., de Vries R.P., Polak S., Seidl V., Seiboth B.

  The Hypocrea jecorina LXR1 was described as the first fungal L-xylulose reductase responsible for NADPH dependent reduction of L-xylulose to xylitol in L-arabinose catabolism. Phylogenetic analysis now reveals that LXR1 forms a clade with fungal D-mannitol 2-dehydrogenases. Lxr1 and the orthologou ... >> More

  The Hypocrea jecorina LXR1 was described as the first fungal L-xylulose reductase responsible for NADPH dependent reduction of L-xylulose to xylitol in L-arabinose catabolism. Phylogenetic analysis now reveals that LXR1 forms a clade with fungal D-mannitol 2-dehydrogenases. Lxr1 and the orthologous Aspergillus niger mtdA are not induced by L-arabinose but expressed at low levels during growth on different carbon sources. Deletion of lxr1 does not affect growth on L-arabinose and L-xylulose reductase activity remains unaltered whereas D-mannitol 2-dehydrogenase activities are reduced. We conclude that LXR1 is a D-mannitol 2-dehydrogenase and that a true LXR1 is still awaiting discovery. << Less

  FEBS Lett. 583:1309-1313(2009) [PubMed] [EuropePMC]

• Yarrowia lipolytica dehydrogenase/reductase: an enzyme tolerant for lipophilic compounds and carbohydrate substrates.

  Napora K., Wrodnigg T.M., Kosmus P., Thonhofer M., Robins K., Winkler M.

  Yarrowia lipolytica short chain dehydrogenase/reductase (YlSDR) was expressed in Escherichia coli, purified and characterized in vitro. The substrate scope for YlSDR mediated oxidation was investigated with alcohols and unprotected carbohydrates spectrophotometrically, revealing a preference for s ... >> More

  Yarrowia lipolytica short chain dehydrogenase/reductase (YlSDR) was expressed in Escherichia coli, purified and characterized in vitro. The substrate scope for YlSDR mediated oxidation was investigated with alcohols and unprotected carbohydrates spectrophotometrically, revealing a preference for secondary compared to primary alcohols. In reduction direction, YlSDR was highly active on ribulose and fructose, suggesting that the enzyme is a mannitol-2-dehydrogenase. In order to explore substrate tolerance especially for space-demanding, lipophilic protecting groups, 5-O-trityl-D-ribitol and 5-O-trityl-α,β-D-ribose were investigated as substrates: YlSDR oxidized 5-O-trityl-D-ribitol and 5-O-trityl-α,β-D-ribose and reduced the latter at the expense of NADP(H). << Less

  Bioorg. Med. Chem. Lett. 23:3393-3395(2013) [PubMed] [EuropePMC]

• Purification and characterization of a novel mannitol dehydrogenase from a newly isolated strain of Candida magnoliae.

  Lee J.K., Koo B.S., Kim S.Y., Hyun H.H.

  Mannitol biosynthesis in Candida magnoliae HH-01 (KCCM-10252), a yeast strain that is currently used for the industrial production of mannitol, is catalyzed by mannitol dehydrogenase (MDH) (EC 1.1.1.138). In this study, NAD(P)H-dependent MDH was purified to homogeneity from C. magnoliae HH-01 by i ... >> More

  Mannitol biosynthesis in Candida magnoliae HH-01 (KCCM-10252), a yeast strain that is currently used for the industrial production of mannitol, is catalyzed by mannitol dehydrogenase (MDH) (EC 1.1.1.138). In this study, NAD(P)H-dependent MDH was purified to homogeneity from C. magnoliae HH-01 by ion-exchange chromatography, hydrophobic interaction chromatography, and affinity chromatography. The relative molecular masses of C. magnoliae MDH, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and size-exclusion chromatography, were 35 and 142 kDa, respectively, indicating that the enzyme is a tetramer. This enzyme catalyzed both fructose reduction and mannitol oxidation. The pH and temperature optima for fructose reduction and mannitol oxidation were 7.5 and 37 degrees C and 10.0 and 40 degrees C, respectively. C. magnoliae MDH showed high substrate specificity and high catalytic efficiency (k(cat) = 823 s(-1), K(m) = 28.0 mM, and k(cat)/K(m) = 29.4 mM(-1) s(-1)) for fructose, which may explain the high mannitol production observed in this strain. Initial velocity and product inhibition studies suggest that the reaction proceeds via a sequential ordered Bi Bi mechanism, and C. magnoliae MDH is specific for transferring the 4-pro-S hydrogen of NADPH, which is typical of a short-chain dehydrogenase reductase (SDR). The internal amino acid sequences of C. magnoliae MDH showed a significant homology with SDRs from various sources, indicating that the C. magnoliae MDH is an NAD(P)H-dependent tetrameric SDR. Although MDHs have been purified and characterized from several other sources, C. magnoliae MDH is distinguished from other MDHs by its high substrate specificity and catalytic efficiency for fructose only, which makes C. magnoliae MDH the ideal choice for industrial applications, including enzymatic synthesis of mannitol and salt-tolerant plants. << Less

  Appl Environ Microbiol 69:4438-4447(2003) [PubMed] [EuropePMC]

• Identification and characterization of the Tuber borchii D-mannitol dehydrogenase which defines a new subfamily within the polyol-specific medium chain dehydrogenases.

  Ceccaroli P., Saltarelli R., Guescini M., Polidori E., Buffalini M., Menotta M., Pierleoni R., Barbieri E., Stocchi V.

  A novel NADP(+)-dependent D-mannitol dehydrogenase and the corresponding gene from the plant symbiotic ascomycete fungus Tuber borchii was identified and characterized. The enzyme, called TbMDH, is a homotetramer with two zinc atoms per subunit. It catalyzed both D-fructose reduction and D-mannito ... >> More

  A novel NADP(+)-dependent D-mannitol dehydrogenase and the corresponding gene from the plant symbiotic ascomycete fungus Tuber borchii was identified and characterized. The enzyme, called TbMDH, is a homotetramer with two zinc atoms per subunit. It catalyzed both D-fructose reduction and D-mannitol oxidation, although it showed the highest substrate specificity and catalytic efficiency for D-fructose. Co-factor specificity was restricted to NADP(H) and the reaction proceeded via a sequential ordered Bi Bi mechanism. The carbon responsive transcriptional pattern showed that Tbmdh is up-regulated when mycelia are transferred to a culture medium containing D-mannitol or D-fructose. The phylogenetic analysis showed TbMDH to be the first example of a fungal D-mannitol-2-dehydrogenase belonging to the medium-chain dehydrogenase/reductases (MDRs). The enzyme identified a new group of proteins, most of them annotated in databases as hypothetical zinc-dependent dehydrogenases, forming a distinct subfamily among the polyol dehydrogenase family. << Less

  Fungal Genet Biol 44:965-978(2007) [PubMed] [EuropePMC]