Publication

The transcarboxylase domain of pyruvate carboxylase is essential for assembly of the peroxisomal flavoenzyme alcohol oxidase

Ozimek, P. Z., Klompmaker, S. H., Visser, N., Veenhuis, M. & van der Klei, I. J., Oct-2007, In : Fems Yeast Research. 7, 7, p. 1082-1092 11 p.

Research output: Contribution to journalArticleAcademicpeer-review

Copy link to clipboard

Documents

DOI

Pyruvate carboxylase (Pyc1p) has multiple functions in methylotrophic yeast species. Besides its function as an enzyme, Pyc1p is required for assembly of peroxisomal alcohol oxidase (AO). Hence, Pyc1p-deficient cells share aspartate auxotrophy (Asp(-)) with a defect in growth on methanol as sole carbon source (Mut(-)). To identify regions in Hansenula polymorpha Pyc1p that are required for the function of HpPyc1p in AO assembly, a series of random mutations was generated in the HpPYC1 gene by transposon mutagenesis. Upon introduction of 18 mutant genes into the H. polymorpha PYC1 deletion strain (pyc1), four different phenotypes were obtained, namely Asp(-) Mut(-), Asp(-) Mut(+), Asp(+) Mut(-), and Asp(+) Mut(+). One mutant showed an Asp(+) Mut(-) phenotype. This mutant produced HpPyc1p containing a pentapeptide insertion in the region that links the conserved N-terminal biotin carboxylation domain (BC) with the central transcarboxylation (TC) domain. Three mutants that were Asp(-) Mut(-) contained insertions in the TC domain, suggesting that this domain is important for both functions of Pyc1p. Analysis of a series of constructed C-terminal and N-terminal truncated versions of HpPyc1p showed that the TC domain of Pyc1p, including the region linking this domain to the BC domain, is essential for AO assembly.

Original languageEnglish
Pages (from-to)1082-1092
Number of pages11
JournalFems Yeast Research
Volume7
Issue number7
Publication statusPublished - Oct-2007
Event4th Hansenula Polymorpha Worldwide Network Conference (HPWN) - , Netherlands
Duration: 3-Sep-20065-Sep-2006

Event

4th Hansenula Polymorpha Worldwide Network Conference (HPWN)

03/09/200605/09/2006

Netherlands

Event: Other

    Keywords

  • pyruvate carboxylase, peroxisome, Hansenula polymorpha, alcohol oxidase, transposon, mutagenesis, FAD, YEAST HANSENULA-POLYMORPHA, PENTAPEPTIDE SCANNING MUTAGENESIS, FLAVIN ADENINE-DINUCLEOTIDE, MOLECULAR CHAPERONES, METHANOL METABOLISM, RECEPTOR PEX5P, PROTEIN, BIOGENESIS, MACHINERY, MECHANISM

Download statistics

No data available

ID: 4612162