High-affinity binding of the yeast cis-Golgi t-SNARE, Sed5p, to wild-type and mutant Sly1p, a modulator of transport vesicle docking
| Autor: | Reiner Grabowski, Dieter Gallwitz |
|---|---|
| Rok vydání: | 1997 |
| Předmět: |
Saccharomyces cerevisiae Proteins
Vesicle docking Recombinant Fusion Proteins Mutant Blotting Western Biophysics Vesicular Transport Proteins Golgi Apparatus Saccharomyces cerevisiae yeast Biology Endoplasmic Reticulum Sly1 protein Biochemistry Fungal Proteins symbols.namesake t-SNARE Munc18 Proteins Sed5 protein Structural Biology Surface plasmon resonance Golgi Genetics Escherichia coli Binding site Cloning Molecular Molecular Biology Binding Sites Qa-SNARE Proteins Vesicle Wild type Membrane Proteins Biological Transport Vesicle transport Cell Biology Golgi apparatus Cell biology Vesicular transport protein Molecular Weight Kinetics Docking (molecular) symbols Chromatography Gel Ypt1 GTPase Electrophoresis Polyacrylamide Gel Carrier Proteins SNARE Proteins Protein Binding |
| Zdroj: | FEBS letters. 411(2-3) |
| ISSN: | 0014-5793 |
| Popis: | Docking of ER-derived vesicles to the cis-Golgi compartment in yeast requires vesicle and target membrane receptors (v-SNAREs and t-SNAREs) and the GTPase Ypt1p. The t-SNARE Sed5p is complexed with Sly1p in vivo. The mutant form Sly1-20p rescues Ypt1p-lacking cells from lethality, suggesting an inhibitory function of Sly1p in v-SNARE/t-SNARE interaction. Using surface plasmon resonance spectroscopy, we found that Sed5p binds Sly1p and Sly1-20p with equally high affinity (K(D) = 5.13 x 10(-9) M and 4.74 x 10(-9) M, respectively). Deletion studies show that the N-terminal half of Sly1p rather than the C-terminus (harbouring the E532K substitution in Sly1-20p) is most critical for its binding to Sed5p. These data appear to argue for an active rather than an inhibitory role of Sly1p in vesicle docking. |
| Databáze: | OpenAIRE |
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