4WNI

Crystal structure of the T229K mutant of human GAPDH at 2.3 angstroems resolution

Summary for 4WNI

• Entry DOI: 10.2210/pdb4wni/pdb
• Related: 4WNC
• Descriptor: Glyceraldehyde-3-phosphate dehydrogenase, NICOTINAMIDE-ADENINE-DINUCLEOTIDE, ZINC ION, ... (4 entities in total)
• Functional Keywords: glycolytic, rossman fold, nad cofactor, mutant, oxidoreductase
• Biological source: Homo sapiens (Human)
• Cellular location: Cytoplasm, cytosol : P04406
• Total number of polymer chains: 4
• Total formula weight: 146695.47

Authors

Garcin, E.D.,White, M.R. (deposition date: 2014-10-12, release date: 2014-12-03, Last modification date: 2023-09-27)

Primary citation

White, M.R.,Khan, M.M.,Deredge, D.,Ross, C.R.,Quintyn, R.,Zucconi, B.E.,Wysocki, V.H.,Wintrode, P.L.,Wilson, G.M.,Garcin, E.D.
A Dimer Interface Mutation in Glyceraldehyde-3-Phosphate Dehydrogenase Regulates Its Binding to AU-rich RNA.
J.Biol.Chem., 290:1770-1785, 2015

PubMed Abstract: 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.

PubMed: 25451934
DOI: 10.1074/jbc.M114.618165

Experimental method

X-RAY DIFFRACTION (2.3 Å)