Penta-EF-hand protein ALG-2 functions as a Ca2+-dependent adaptor that bridges Alix and TSG101
| Autor: | Haruna Yoshida, Hironori Suzuki, Mayumi Okumura, Ryota Kobayashi, Fumitaka Ichioka, Hideki Shibata, Masatoshi Maki |
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| Rok vydání: | 2009 |
| Předmět: |
Gene isoform
Mutant Biophysics Cell Cycle Proteins macromolecular substances Biology Crystallography X-Ray Biochemistry Protein–protein interaction Cell Line Calcium-binding protein TSG101 Humans Protein Isoforms Molecular Biology Endosomal Sorting Complexes Required for Transport EF hand HEK 293 cells Calcium-Binding Proteins Signal transducing adaptor protein Cell Biology Molecular biology Cell biology DNA-Binding Proteins Calcium Apoptosis Regulatory Proteins Transcription Factors |
| Zdroj: | Biochemical and biophysical research communications. 386(1) |
| ISSN: | 1090-2104 |
| Popis: | Alix and TSG101, known to physically interact with each other, have Pro-rich regions that are bound by ALG-2 Ca2+-dependently. We investigated the role of ALG-2 in the Alix-TSG101 association by pulldown assays using Strep-tagged Alix and its various mutants. The ALG-2-binding site was required for the Ca2+-dependent pulldown of TSG101 using HEK293T cells, whereas the PSAP sequence, a binding motif for the UEV domain of TSG101, was dispensable. Alix-TSG101 association was not observed using ALG-2-knockdown cells but became detectable by addition of the purified recombinant ALG-2 protein in the assay mixtures. Exogenous expression of mGFP-fused ALG-2 also restored the pulldown capability of Strep-Alix, but an alternatively spliced shorter ALG-2 isoform and a dimerization-defective mutant were incompetent. Based on the X-ray crystal structure model showing the presence of one ligand-binding site in each molecule of an ALG-2 dimer, we propose that Ca2+-loaded ALG-2 bridges Alix and TSG101 as an adaptor protein. |
| Databáze: | OpenAIRE |
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