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In vivo processing of nonanchored Yapsin 1 (Yap3p)
1Section on Cellular Neurobiology, Laboratory of Developmental Neurobiology, National Institute of Child Health and Human Development, Building 49, Room 5A38, Bethesda, Maryland, 20892, USA. ypl@codon.nih.gov
Archives of Biochemistry and Biophysics
|March 4, 2000
Summary
Yeast aspartic protease 3 (yapsin 1) processing was studied. The 87 kDa pro-yapsin 1 is autocatalytically cleaved in the endoplasmic reticulum into 18 kDa alpha and 74 kDa beta subunits.
Area of Science:
- Biochemistry
- Molecular Biology
- Yeast Genetics
Background:
- Yapsin 1, a yeast aspartic protease, plays a role in cellular processes.
- Understanding protein processing is crucial for deciphering enzyme function.
Purpose of the Study:
- To investigate the processing pathway of a C-terminally truncated yapsin 1 in yeast.
- To identify the molecular mechanisms and cellular compartments involved in yapsin 1 maturation.
Main Methods:
- Overexpression of truncated yapsin 1 in Saccharomyces cerevisiae.
- Pulse-labeling and immunoprecipitation to track protein processing.
- Utilizing a temperature-sensitive sec18 mutant strain to determine the site of processing.
Main Results:
- Three forms of yapsin 1 were identified: 87 kDa (pro-yapsin 1), 74 kDa (beta subunit), and 18 kDa (alpha subunit).
- Processing into alpha and beta subunits occurs via cleavage at a unique loop region.
- Subunit generation was localized to the endoplasmic reticulum using the sec18 mutant.
- An active site mutation prevented subunit generation, indicating autocatalytic processing.
Conclusions:
- Yapsin 1 undergoes autocatalytic processing in the endoplasmic reticulum.
- The cleavage mechanism differs from traditional aspartic proteases.
- This study elucidates a novel protein maturation pathway in yeast.