N-terminal entrance loop of yeast Yps1 and O-glycosylation of substrates are determinant factors controlling the shedding activity of this GPI-anchored endopeptidase
Bibliographic record
Abstract
BACKGROUND: S. cerevisiae Yps1 is the prototypical aspartic endopeptidase of the fungal yapsin family. This glycosylphosphatidylinositol (GPI) anchored enzyme was recently shown to be involved in the shedding of the GPI proteins Utr2, Gas1 and itself. It was also proposed to be part of a novel quality control mechanism that eliminates excess and/or misfolded GPI proteins. What regulates its shedding activity at the cell surface is however poorly understood. Yps1 is initially synthesized as a zymogen requiring proteolytic activation to remove a pro-peptide and further processing within a large insertion loop (N-entrance loop) generates a two-subunit endopeptidase. To investigate the role of this loop on its shedding activity, which typically takes place within Ser/Thr-rich domains, it was replaced with the short peptide found at the analogous position in Yps3. We also tested whether O-glycosylation might protect against proteolytic processing by Yps1. RESULTS: We show here that replacement of the N-entrance loop (N-ent loop) of Yps1 generates a single chain endopeptidase that undergoes partial (pH 6.0) or complete (pH 3.0) pro-peptide removal. At both pH, the shedding activity of the chimeric endopeptidase (Yps1-DL) toward Gas1 and itself is strongly and drastically increased, respectively. A direct correlation between endoproteolytic cleavage of this loop in native Yps1 and its shedding is observed. The Yps1-dependent shedding of two model GPI proteins (Gas1 and Yps1) is also stimulated by the absence of the O-mannosyltransferases, Pmt4 and Pmt2 respectively, involved in O-glycosylation of their Ser/Thr-rich domains. Under these conditions, some Yps1-independent shedding is also observed. CONCLUSIONS: Partial pro-peptide removal is essential to produce a functional Yps1 endopeptidase. The Yps1 N-ent loop plays a major role in regulating the shedding activity of the endopeptidase, most likely by limiting access to the active site, and its cleavage in native Yps1 is associated with its shedding. O-glycosylation protects against Yps1-dependent and -independent shedding of GPI proteins. It is postulated that hypoglycosylation of cell surface proteins, which may occur for misfolded proteins that escaped the ER-associated degradation, might target their elimination through shedding by Yps1 and possibly other yapsin members.
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".