Regulation of methanol metabolism in the yeast Hansenula polymorpha

Koning, W. de; Gleeson, Martin A.; Harder, W.; Dijkhuizen, Lubbert

doi:10.1007/bf00406136

Cited by 30 publications

(9 citation statements)

References 21 publications

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“…The DAS synthesis in glycerol-grown cells points to the idea that formaldehyde is not indispensable for the enzyme induction, and a mechanism for DAS derepression in H. polymorpha has been proposed [16]. However, the latter has not been proved by other authors [17,18] or by our results.…”

Section: Resultscontrasting

confidence: 54%

Methanol metabolism in mutants of the methylotrophic yeast Hansenula polymorpha

Bystrykh¹

1988

FEMS Microbiology Letters

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Section: Resultscontrasting

confidence: 54%

Methanol metabolism in mutants of the methylotrophic yeast Hansenula polymorpha

Bystrykh¹

1988

FEMS Microbiology Letters

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“…This is not due to the presence of an isoform of DAK because no homologous gene could be identified by Southern blot analysis and no enzymatic activity was present in the dak mutant strain, implicating an alternative but less efficient pathway for DHA metabolism. A similar phenotype has been observed for a H. polymorpha mutant (17B) lacking DAK activity (de Koning et al, 1987a). Using a double mutant lacking DAK and dihydroxyacetone synthase activities, it was shown (de Koning et al, 1987a) that the growth rate of the Hpdak-deficient strain 17B on DHA was not due to the reverse dihydroxyacetone synthase reaction.…”

Section: Discussionsupporting

confidence: 66%

“…A similar phenotype has been observed for a H. polymorpha mutant (17B) lacking DAK activity (de Koning et al, 1987a). Using a double mutant lacking DAK and dihydroxyacetone synthase activities, it was shown (de Koning et al, 1987a) that the growth rate of the Hpdak-deficient strain 17B on DHA was not due to the reverse dihydroxyacetone synthase reaction. The slow growth rate was abolished in double mutants lacking DAK activity and either glycerol kinase or glycerol-3-phosphate dehydrogenase activity.…”

Section: Discussionsupporting

confidence: 66%

ThePichia pastoris dihydroxyacetone kinase is a PTS1-containing, but cytosolic, protein that is essential for growth on methanol

Lüers

Advani

Wenzel

et al. 1998

Yeast

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Dihydroxyacetone kinase (DAK) is essential for methanol assimilation in methylotrophic yeasts. We have cloned the DAK gene from Pichia pastoris by functional complementation of a mutant that was unable to grow on methanol. An open reading frame of 1824 bp was identified that encodes a 65·3 kDa protein with high homology to DAK from Saccharomyces cerevisiae. Although DAK from P. pastoris contained a C‐terminal tripeptide, TKL, which we showed can act as a peroxisomal targeting signal when fused to the green fluorescent protein, the enzyme was primarily cytosolic. The TKL tripeptide was not required for the biochemical function of DAK because a deletion construct lacking the DNA encoding this tripeptide was able to complement the P. pastoris dakΔ mutant. Peroxisomes, which are essential for growth of P. pastoris on methanol, were present in the dakΔ mutant and the import of peroxisomal proteins was not disturbed. The dakΔ mutant grew at normal rates on glycerol and oleate media. However, unlike the wild‐type cells, the dakΔ mutant was unable to grow on methanol as the sole carbon source but was able to grow on dihydroxyacetone at a much slower rate. The metabolic pathway explaining the reduced growth rate of the dakΔ mutant on dihydroxyacetone is discussed. The nucleotide sequence reported in this paper has been submitted to GenBank with Accession Number AF019198. © 1998 John Wiley & Sons, Ltd.

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“…Cell free extracts for determination of enzyme activities were prepared as described by De Koning et al (1987). Enzyme assays were carried out at 37°C.…”

Section: Cell-free Extract Preparation and Enzyme Assaysmentioning

confidence: 99%

Alcohol oxidase expressed under nonmethylotrophic conditions is imported, assembled, and enzymatically active in peroxisomes of Hansenula polymorpha.

Distel

Ley

Veenhuis

et al. 1988

The Journal of Cell Biology

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Abstract. We have introduced into Hansenula polymorpha an extra copy of its alcohol oxidase gene. This gene which is under the control of the Saccharomyces cerevisiae phosphoglycerate kinase promoter is integrated in a chromosome different from the one conmining the endogenous gene. Cells with the extra alcohol oxidase gene, grown on glucose or ethanol as the sole carbon source, express enzymatically active alcohol oxidase. However, other enzymes characteristic for methylotrophic growth conditions are absent or present at low levels. Most of the alcohol oxidase occurs in the octameric state and immuno-and cytochemical evidence shows that it is located in a single enlarged peroxisome per cell. Such peroxisomes show crystalloid inclusions which are lacking in the peroxisomes present in glucose grown control cells.Our results suggest that import into peroxisomes of H. polymorpha, assembly and activation of alcohol oxidase is not conditionally dependent on adaptation to methylotrophic growth conditions and that proliferation of peroxisomes is a well-programmed process that is not triggered solely by overproduction of a peroxisomal protein.LIKARYOTIC cells contain a number of membrane-enclosed compartments each devoted to carry out specific metabolic functions. To this purpose each organelle possesses its own characteristic group of enzymes and proteins. Some of these organeUes have been favorite objects of research and much is already known about their contribution to cellular metabolism and biogenesis. Relatively little is known however, about peroxisomes, which were first discovered by De Duve et al. (1966). Although rather variable in enzymatic repertoire depending on the organism or tissue in which they occur, a common feature is the presence of an H202 producing oxidase and catalase. Recent studies have revealed that peroxisomal proteins, including integral membrane proteins, are encoded in the nucleus, synthesized on free polysomes and imported posttranslationally into the organelle (reviewed by Lazarow and Fujiki, 1985). Moreover, the majority of peroxisomal proteins are synthesized at their mature size and translocation across the peroxisomal membrane occurs without any detectable modification of the protein (reviewed by Borst, 1986).A very interesting example of peroxisomal function and development is represented by methylotrophic yeasts. In these fungi the number of organelles and their physiological function is entirely dependent upon environmental conditions (for reviews see Veenhuis et al., 1983 and. For instance when the methylotrophic yeast Hansenula polymorpha is grown on methanol the cells contain a large number of peroxisomes which have a crystalline matrix exclusively composed of alcohol oxidase (AO)~ molecules (Veenhuis et al., 1978;Veenhuis et al., 1981). AO functions as the first enzyme in methanol metabolism converting methanol to formaldehyde and H202. Besides AO considerable amounts of catalase and dihydroxy acetone synthase are present in these organelles (Douma et al., 1985;Goodman, 1985)....

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Regulation of methanol metabolism in the yeast Hansenula polymorpha

Cited by 30 publications

References 21 publications

Methanol metabolism in mutants of the methylotrophic yeast Hansenula polymorpha

Methanol metabolism in mutants of the methylotrophic yeast Hansenula polymorpha

ThePichia pastoris dihydroxyacetone kinase is a PTS1-containing, but cytosolic, protein that is essential for growth on methanol

Alcohol oxidase expressed under nonmethylotrophic conditions is imported, assembled, and enzymatically active in peroxisomes of Hansenula polymorpha.

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