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Characterization of the Hansenula polymorpha CPY gene encoding carboxypeptidase Y
A R Bellu1, I J van der Klei, K B Rechinger
1Eukaryotic Microbiology, Griningen Biomolecular Sciences and Biotechnology Institute (GBB), University of Groningen, Haren, The Netherlands.
Yeast (Chichester, England)
|March 17, 1999
Summary
We isolated the Hansenula polymorpha carboxypeptidase Y (Hp-CPY) gene. Deleting this gene did not affect peroxisome degradation, indicating Hp-CPY is not essential for this autophagous process.
Area of Science:
- Molecular biology
- Cell biology
- Yeast genetics
Background:
- Carboxypeptidase Y (CPY) is a key vacuolar enzyme.
- Peroxisome degradation is an essential autophagous process in yeast.
- Understanding CPY's role in yeast cellular processes is important.
Purpose of the Study:
- To isolate and characterize the Hansenula polymorpha CPY gene (Hp-CPY).
- To investigate the role of Hp-CPY in peroxisome degradation.
Main Methods:
- Gene isolation and sequencing of Hp-CPY.
- Amino acid sequence analysis and comparison with Saccharomyces cerevisiae CPY.
- Immunocytochemistry and Western blotting to localize and detect Hp-CPY.
- Endoglycosidase H treatment to assess glycosylation.
- Analysis of peroxisome degradation in deltacpy mutant cells.
Main Results:
- The Hp-CPY gene encodes a 541-amino acid protein with high similarity to S. cerevisiae CPY.
- Hp-CPY contains signals for secretion and vacuolar sorting.
- Hp-CPY is localized to the vacuole and is glycosylated.
- Deletion of the Hp-CPY gene did not impair the degradation of peroxisomal proteins.
Conclusions:
- Hp-CPY is a functional carboxypeptidase Y in Hansenula polymorpha.
- Hp-CPY is not a major proteinase involved in the autophagous degradation of peroxisomes.