TY - JOUR
T1 - Mechanisms for ligand binding to GluR0 ion channels
T2 - Crystal structures of the glutamate and serine complexes and a closed apo state
AU - Mayer, Mark L.
AU - Olson, Rich
AU - Gouaux, Eric
N1 - Funding Information:
We thank N. Armstrong, E. Braswell, G. Q. Chen and K. Fleming for advice; N. Armstrong and D. Yernool for help with data collection; J. Lidestri for maintenance of the X-ray facility; C. Ogata and R. Abramowitz for support at beamline X4A, National Synchrotron Light Source; Y. Jin for plasmid construction; M. Gawinowicz for performing mass spectral analysis and protein sequencing; N. Armstrong and C. Cui for comments on the manuscript. E.G. is an assistant investigator of the Howard Hughes Medical Institute and research in his laboratory is also supported by grants from the NIH. The XLI ultracentrifuge was obtained from funds provided by an NIH shared instrumentation grant (S10 RR12848) and the R-Axis 4 X-ray diffraction system was purchased using funds from the NSF and Columbia University. M.L.M. was supported by the NICHD intramural program while on sabbatical leave at Columbia University and thanks D. Hirsh, the faculty and graduate students of the Department of Biochemistry and Molecular Biophysics for numerous acts of kindness.
PY - 2001/8/24
Y1 - 2001/8/24
N2 - High-resolution structures of the ligand binding core of GluR0, a glutamate receptor ion channel from Synechocystis PCC 6803, have been solved by X-ray diffraction. The GluR0 structures reveal homology with bacterial periplasmic binding proteins and the rat GluR2 AMPA subtype neurotransmitter receptor. The ligand binding site is formed by a cleft between two globular α/β domains. L-Glutamate binds in an extended conformation, similar to that observed for glutamine binding protein (GlnBP). However, the L-glutamate γ-carboxyl group interacts exclusively with Asn51 in domain 1, different from the interactions of ligand with domain 2 residues observed for GluR2 and GlnBP. To address how neutral amino acids activate GluR0 gating we solved the structure of the binding site complex with L-serine. This revealed solvent molecules acting as surrogate ligand atoms, such that the serine OH group makes solvent-mediated hydrogen bonds with Asn51. The structure of a ligand-free, closed-cleft conformation revealed an extensive hydrogen bond network mediated by solvent molecules. Equilibrium centrifugation analysis revealed dimerization of the GluR0 ligand binding core with a dissociation constant of 0.8 μM. In the crystal, a symmetrical dimer involving residues in domain 1 occurs along a crystallographic 2-fold axis and suggests that tetrameric glutamate receptor ion channels are assembled from dimers of dimers. We propose that ligand-induced conformational changes cause the ion channel to open as a result of an increase in domain 2 separation relative to the dimer interface.
AB - High-resolution structures of the ligand binding core of GluR0, a glutamate receptor ion channel from Synechocystis PCC 6803, have been solved by X-ray diffraction. The GluR0 structures reveal homology with bacterial periplasmic binding proteins and the rat GluR2 AMPA subtype neurotransmitter receptor. The ligand binding site is formed by a cleft between two globular α/β domains. L-Glutamate binds in an extended conformation, similar to that observed for glutamine binding protein (GlnBP). However, the L-glutamate γ-carboxyl group interacts exclusively with Asn51 in domain 1, different from the interactions of ligand with domain 2 residues observed for GluR2 and GlnBP. To address how neutral amino acids activate GluR0 gating we solved the structure of the binding site complex with L-serine. This revealed solvent molecules acting as surrogate ligand atoms, such that the serine OH group makes solvent-mediated hydrogen bonds with Asn51. The structure of a ligand-free, closed-cleft conformation revealed an extensive hydrogen bond network mediated by solvent molecules. Equilibrium centrifugation analysis revealed dimerization of the GluR0 ligand binding core with a dissociation constant of 0.8 μM. In the crystal, a symmetrical dimer involving residues in domain 1 occurs along a crystallographic 2-fold axis and suggests that tetrameric glutamate receptor ion channels are assembled from dimers of dimers. We propose that ligand-induced conformational changes cause the ion channel to open as a result of an increase in domain 2 separation relative to the dimer interface.
KW - Equilibrium centrifugation
KW - Glutamate receptors
KW - Ion channels
KW - Ligand binding
KW - X-ray crystallography
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U2 - 10.1006/jmbi.2001.4884
DO - 10.1006/jmbi.2001.4884
M3 - Article
C2 - 11518533
AN - SCOPUS:0035943408
SN - 0022-2836
VL - 311
SP - 815
EP - 836
JO - Journal of molecular biology
JF - Journal of molecular biology
IS - 4
ER -