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Physiological role of the glutathione-dependent formaldehyde dehydrogenase in the methylotrophic yeast Candida boidinii^b

Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-Oiwake, Sakyo-ku, Kyoto 606-8502, Japan¹

Author for correspondence: Yasuyoshi Sakai. Tel: +81 75 753 6455. Fax: +81 75 753 6385. e-mail: ysakai{at}kais.kyoto-u.ac.jp

The methylotrophic yeast Candida boidinii exhibits formaldehyde dehydrogenase activity (FLD, EC 1.2.1.1) during growth on methanol as a sole carbon source. The structural gene, FLD1, was cloned from a genomic library of C. boidinii. The 1263 bp FLD1 gene contained a 123 bp intron and its exon encoded a gene product of 380 amino acids, whose predicted amino acid sequence showed high similarity to the sequences of FLDs from other organisms. The FLD1 gene was disrupted in the C. boidinii genome by one-step gene disruption. The fld1 strain could not grow on methanol as a carbon source under methanol-limited chemostat culture conditions, even with low dilution rates (D<0·05 h^-1), whereas a strain with a disruption in the gene for formate dehydrogenase (FDH; another NADH-generating dehydrogenase involved in the formaldehyde oxidation pathway) could survive. These results indicated that FLD, but not FDH, is essential for growth of C. boidinii on methanol.

Keywords: formaldehyde oxidation pathway, gene cloning, gene disruption, energy generation, formaldehyde detoxification

Abbreviations: AOD, alcohol oxidase; DHAS, dihydroxyacetone synthase; FDH, formate dehydrogenase; FLD, glutathione-dependent formaldehyde dehydrogenase

^b The GenBank accession number for the sequence reported in this paper is AB085186.

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