The Hansenula polymorpha per6 mutant is affected in two adjacent genes which encode dihydroxyacetone kinase and a novel protein, Pak1p, involved in peroxisome integrity

Abstract

The Hansenula polymorpha per6 mutant is affected in two adjacent genes which encode dihydroxyacetone kinase and a novel protein, Pak1p, involved in peroxisome integrity Klei, Ida J. van der; Heide, Meis van der; Baerends, Richard J.S.; Rechinger, Karl-Björn; Nicolay, Klaas; Kiel, Jan A.K.W.; Veenhuis, Marten Take-down policy If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim.Downloaded from the University of Groningen/UMCG research database (Pure): http://www.rug.nl/research/portal. For technical reasons the number of authors shown on this cover page is limited to 10 maximum. Download date: 04-04-2019Abstract The Hansenula polymorpha per6-210 mutant is impaired in respect of growth on methanol (Mut -) and is characterized by aberrant peroxisome formation. The functionally complementing DNA fragment contains two open reading frames. The first encodes dihydroxyacetone kinase (DAK), a cytosolic enzyme essential for formaldehyde assimilation; the second ORF codes for a novel protein (Pak1p). We have demonstrated that per6-210 cells lack DAK activity, causing the Mut -phenotype, and have strongly reduced levels of Pak1p, resulting in peroxisomal defects. Sequence analysis revealed that per6-210 contains a mutation in the 3′ end of the DAK coding region, which overlaps with the promoter region of PAK1. Possibly this mutation also negatively affects PAK1 expression.

Cloning, sequencing and characterization of a gene encoding dihydroxyacetone kinase from Zygosaccharomyces rouxii NRRL2547

“…Whereas two DAK genes are found in S. cerevisiae (Norbeck and Blomberg, 1997) and Sz. pombe (Kimura et al, 1998), only one DAK gene has been reported in other yeasts, such as Pichia pastoris and Pichia angusta (Luers et al, 1998;van der Klei et al, 1998). In this study, we failed to detect any other DAK genes in Z. rouxii, even under non-stringent hybridization conditions.…”

“…Although many yeasts apparently use this so-called G3P pathway to metabolize glycerol (Prior and Hohmann, 1997), an alternative metabolic pathway, known as the DHA pathway, for glycerol dissimilation that requires a glycerol dehydrogenase and a dihydroxyacetone kinase has been found in some yeast species. These include Schizosaccharomyces pombe (May and Sloan, 1981;Gancedo et al, 1986), Debaryomyces hansenii (Adler et al, 1985), S. cerevisiae (Babel and Hofmann, 1982), Zygosaccharomyces rouxii (van Zyl et al, 1991) and species of Candida (Hofmann and Babel, 1980), Hansenula and Pichia (Tikhomirova et al, 1988;Luers et al, 1998;van der Klei et al, 1998). In S. cerevisiae, the genes GCY1 and DAK1, encoding a putative glycerol dehydrogenase (EC 1.1.1.6) and dihydroxyacetone kinase (EC 2.7.1.29), respectively, have been cloned and partially characterized (Norbeck and Blomberg, 1997; Molin M, Norbeck J, Blomberg A, submitted for publication).…”

Hansenula polymorpha Pex3p Is a Peripheral Component of the Peroxisomal Membrane

“…Biochemical Methods-Crude extracts (18), protoplasts (19), and organellar fractions (20) were prepared as described. Protease inhibitors were omitted when organellar fractions were used for protease protection assays.…”

Hansenula polymorphaPex20p is an oligomer that binds the peroxisomal targeting signal 2 (PTS2)

“…Cell fractionation studies were performed as detailed previously (van der Klei et al, 1998), except that 1 mM NaF and 1 mM PMSF was added to all buffers.…”