A dimer interface mutation in glyceraldehyde 3-phosphate dehydrogenase regulates its binding to AU-rich RNA
| Autor: | Christina R. Ross, Beth E. Zucconi, Vicki H. Wysocki, Daniel Deredge, Michael R. White, Mohd M. Khan, Royston S. Quintyn, Patrick L. Wintrode, Elsa D. Garcin, Gerald M. Wilson |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2015 |
| Předmět: |
Spectrometry
Mass Electrospray Ionization DNA Complementary Protein Conformation Amino Acid Motifs Molecular Sequence Data Biochemistry stomatognathic system Tetramer Oxidoreductase Humans Scattering Radiation Amino Acid Sequence Nucleic acid structure Binding site Molecular Biology 3' Untranslated Regions Glyceraldehyde 3-phosphate dehydrogenase AU-rich element chemistry.chemical_classification Messenger RNA Binding Sites biology Sequence Homology Amino Acid Tumor Necrosis Factor-alpha X-Rays RNA Glyceraldehyde-3-Phosphate Dehydrogenases Cell Biology chemistry Microscopy Fluorescence Mutation biology.protein Anisotropy Additions and Corrections Protein Multimerization Peptides Glycolysis Molecular Biophysics Protein Binding |
| Popis: | Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is an enzyme best known for its role in glycolysis. However, extra-glycolytic functions of GAPDH have been described, including regulation of protein expression via RNA binding. GAPDH binds to numerous adenine-uridine rich elements (AREs) from various mRNA 3'-untranslated regions in vitro and in vivo despite its lack of a canonical RNA binding motif. How GAPDH binds to these AREs is still unknown. Here we discovered that GAPDH binds with high affinity to the core ARE from tumor necrosis factor-α mRNA via a two-step binding mechanism. We demonstrate that a mutation at the GAPDH dimer interface impairs formation of the second RNA-GAPDH complex and leads to changes in the RNA structure. We investigated the effect of this interfacial mutation on GAPDH oligomerization by crystallography, small-angle x-ray scattering, nano-electrospray ionization native mass spectrometry, and hydrogen-deuterium exchange mass spectrometry. We show that the mutation does not significantly affect GAPDH tetramerization as previously proposed. Instead, the mutation promotes short-range and long-range dynamic changes in regions located at the dimer and tetramer interface and in the NAD(+) binding site. These dynamic changes are localized along the P axis of the GAPDH tetramer, suggesting that this region is important for RNA binding. Based on our results, we propose a model for sequential GAPDH binding to RNA via residues located at the dimer and tetramer interfaces. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|