Abstract
In yeast the key gluconeogenic enzyme, fructose-1,6-bisphosphatase (FBPase), is selectively targeted from the cytosol to the lysosome (vacuole) for degradation when glucose starved cells are replenished with glucose. The pathway for glucose induced FBPase degradation is unknown. To identify the receptor-mediated degradation pathway of FBPase, we investigated the presence of the FBPase receptor on the vacuolar membrane by cell fractionation experiments and binding assay using rid mutant (vacuolar import and degradation), which is defective in the glucose-induced degradation of FBPase. FBPase sedimented in the pellets from vid24-1 mutant after centrifugation at 15,000 × g for 15 min, suggesting that FBPase is associated with subcellular structures. Cell fractionation experiments revealed that FBPase is preferentially associated with the vacuole, but not with other organelles in vid24-1. FBPase enriched fractions that co-fractionated with the vacuole were sensitive to proteinase K digestion, indicating that FBPase is peripherally associated with the vacuole. We developed an assay for the binding of FBPase to the vacuole. The assay revealed that FBPase bound to the vacuole with a Kd of 2.3 × 106M. The binding was saturable and specific. These results suggest that a receptor for FBPase degradation exists on the vacuolar membrane. It implies the existence of the receptor-mediated degradation pathway of FBPase by the lysosome.
| Original language | English |
|---|---|
| Pages (from-to) | 448-453 |
| Number of pages | 6 |
| Journal | Journal of Biochemistry and Molecular Biology |
| Volume | 33 |
| Issue number | 6 |
| Publication status | Published - 2000 Nov 30 |
| Externally published | Yes |
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Keywords
- FBPase receptor
- Fructose-1,6-bisphosphatase
- Protein degradation
- Vacuolar degradation pathway
- Vid24-1 mutant
ASJC Scopus subject areas
- Biochemistry
- Molecular Biology
Cite this
Identification of Receptor-like Protein for Fructose-1,6-bisphosphatase on Yeast Vacuolar Membrane. / Ko, Je Sang.
In: Journal of Biochemistry and Molecular Biology, Vol. 33, No. 6, 30.11.2000, p. 448-453.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Identification of Receptor-like Protein for Fructose-1,6-bisphosphatase on Yeast Vacuolar Membrane
AU - Ko, Je Sang
PY - 2000/11/30
Y1 - 2000/11/30
N2 - In yeast the key gluconeogenic enzyme, fructose-1,6-bisphosphatase (FBPase), is selectively targeted from the cytosol to the lysosome (vacuole) for degradation when glucose starved cells are replenished with glucose. The pathway for glucose induced FBPase degradation is unknown. To identify the receptor-mediated degradation pathway of FBPase, we investigated the presence of the FBPase receptor on the vacuolar membrane by cell fractionation experiments and binding assay using rid mutant (vacuolar import and degradation), which is defective in the glucose-induced degradation of FBPase. FBPase sedimented in the pellets from vid24-1 mutant after centrifugation at 15,000 × g for 15 min, suggesting that FBPase is associated with subcellular structures. Cell fractionation experiments revealed that FBPase is preferentially associated with the vacuole, but not with other organelles in vid24-1. FBPase enriched fractions that co-fractionated with the vacuole were sensitive to proteinase K digestion, indicating that FBPase is peripherally associated with the vacuole. We developed an assay for the binding of FBPase to the vacuole. The assay revealed that FBPase bound to the vacuole with a Kd of 2.3 × 106M. The binding was saturable and specific. These results suggest that a receptor for FBPase degradation exists on the vacuolar membrane. It implies the existence of the receptor-mediated degradation pathway of FBPase by the lysosome.
AB - In yeast the key gluconeogenic enzyme, fructose-1,6-bisphosphatase (FBPase), is selectively targeted from the cytosol to the lysosome (vacuole) for degradation when glucose starved cells are replenished with glucose. The pathway for glucose induced FBPase degradation is unknown. To identify the receptor-mediated degradation pathway of FBPase, we investigated the presence of the FBPase receptor on the vacuolar membrane by cell fractionation experiments and binding assay using rid mutant (vacuolar import and degradation), which is defective in the glucose-induced degradation of FBPase. FBPase sedimented in the pellets from vid24-1 mutant after centrifugation at 15,000 × g for 15 min, suggesting that FBPase is associated with subcellular structures. Cell fractionation experiments revealed that FBPase is preferentially associated with the vacuole, but not with other organelles in vid24-1. FBPase enriched fractions that co-fractionated with the vacuole were sensitive to proteinase K digestion, indicating that FBPase is peripherally associated with the vacuole. We developed an assay for the binding of FBPase to the vacuole. The assay revealed that FBPase bound to the vacuole with a Kd of 2.3 × 106M. The binding was saturable and specific. These results suggest that a receptor for FBPase degradation exists on the vacuolar membrane. It implies the existence of the receptor-mediated degradation pathway of FBPase by the lysosome.
KW - FBPase receptor
KW - Fructose-1,6-bisphosphatase
KW - Protein degradation
KW - Vacuolar degradation pathway
KW - Vid24-1 mutant
UR - http://www.scopus.com/inward/record.url?scp=0034357245&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0034357245&partnerID=8YFLogxK
M3 - Article
AN - SCOPUS:0034357245
VL - 33
SP - 448
EP - 453
JO - BMB Reports
JF - BMB Reports
SN - 1976-6696
IS - 6
ER -