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Applied and Environmental Microbiology, December 1998, p. 4689-4696, Vol. 64, No. 12
0099-2240/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Cloning and Characterization of NADP-Mannitol Dehydrogenase cDNA from the Button Mushroom, Agaricus bisporus, and Its Expression in Response to NaCl Stress

Johan M. H. Stoop and Hans Mooibroek*

Department of Industrial Agrobiotechnology, Agrotechnological Research Institute (ATO-DLO), NL-6700 AA Wageningen, The Netherlands

Received 8 July 1998/Accepted 15 September 1998

Mannitol, a six-carbon sugar alcohol, is the main storage carbon in the button mushroom, Agaricus bisporus. Given the physiological importance of mannitol metabolism in growth, fruit body development, and salt tolerance of A. bisporus, the enzyme responsible for mannitol biosynthesis, NADP-dependent mannitol dehydrogenase (MtDH) (EC 1.1.1.138), was purified to homogeneity, and MtDH cDNA was cloned, sequenced, and characterized. To our knowledge, this represents the first report on the isolation of a cDNA encoding an NADP-dependent mannitol dehydrogenase. The MtDH cDNA contains an open reading frame of 789 bp encoding a protein of approximately 28 kDa. The N-terminal and internal amino acid sequences of the deduced protein exactly matched the ones determined from the purified MtDH subunit, whereas the amino acid composition of the deduced protein was nearly identical to that of the purified MtDH. The MtDH cDNA showed high homology with a plant-induced short-chain dehydrogenase from Uromyces fabae. Phylogenetic analysis based on amino acid sequences from mannitol(-1-phosphate) dehydrogenases indicated a close relationship between the substrate specificity of the enzymes and phylogenetic differentiation. Salt-stressed fruit bodies showed an overall increase in mannitol biosynthesis, as was evident from the increase in MtDH activity, MtDH abundance, and MtDH RNA accumulation. Furthermore, the MtDH transcript level seems to be under developmental control, as MtDH RNA accumulated during maturation of the fruit body.

* Corresponding author. Mailing address: Agrotechnological Research Institute (ATO-DLO), Bornsesteeg 59, P.O. Box 17, NL-6700 AA Wageningen, The Netherlands. Phone: (31) 317 47 53 16. Fax: (31) 317 47 53 47. E-mail: a.mooibroek{at}ato.dlo.nl.

Applied and Environmental Microbiology, December 1998, p. 4689-4696, Vol. 64, No. 12
0099-2240/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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