Identification and Mechanism of Action of Two Histidine Residues Underlying High-Affinity Zn2+ Inhibition of the NMDA Receptor

Choi, Yun-Beom; Lipton, Stuart A.

doi:10.1016/s0896-6273(00)80763-1

Cited by 167 publications

(195 citation statements)

References 44 publications

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“…This conclusion is consistent with the findings that the PSD contains several zinc-binding proteins, e.g. protein kinase C (Hubbard et al 1991), dystrophin (Kim et al 1992), NMDA receptors (Choi and Lipton 1999), SAP-102 (El-Husseini et al 2000) and tubulin (Hesketh 1983). In this study, we further found that treatment with 8 M urea resulted in the depletion of most of the zinc ions bound to the PSDs and, concomitantly, disruption of the PSD structure.…”

Section: Discussionsupporting

confidence: 93%

Structural role of zinc ions bound to postsynaptic densities

Jan

Chen

Tsai

et al. 2002

Journal of Neurochemistry

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Postsynaptic densities (PSDs) isolated from porcine cerebral cortices are large aggregates consisting of more than 30 different proteins. Inductively coupled plasma-mass spectrometric analyses revealed that isolated PSDs contained zinc at a concentration of 4.1 nmol per mg protein. Treatment with 8 M urea lead to dissociation of the PSDs into small components and, concomitantly, depletion of most of their bound zinc. After removal of the urea by dialysis, urea-dissociated PSD proteins did not reassemble into aggregates by themselves. Adding ZnCl 2 to urea-treated PSD samples resulted in the assembly of urea-dissociated proteins into large aggregates with morphology and protein composition closely resembling those of the original PSDs. Mg 2+ , Ca 2+ , Co 2+ , Cd 2+ , Cu 2+ , Mn 2+ , Fe 3+ , K + and Na + ions at higher concentrations also induced the aggregation of urea-dissociated PSD protein. The structures of the K + -, Na + -, Mg 2+ -and Ca 2+ -induced aggregates were distinct from that of the original PSDs. Our results indicate that the structure of the PSD could be disassembled and reassembled under in vitro conditions. They further suggest that Zn 2+ ions, by binding to certain zincbinding proteins, play an important role in the formation and maintenance of the structure of the PSD.

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Section: Discussionsupporting

confidence: 93%

Structural role of zinc ions bound to postsynaptic densities

Jan

Chen

Tsai

et al. 2002

Journal of Neurochemistry

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“…The finding that protons appear to shift receptors to an inactive state also carries structural implications and suggests that other forms of allosteric modulation could proceed through creation of briefly occupied inactive states [e.g., calcineurin effects on GluR1 (Banke et al, 2000)]. Given that a number of regulators, such as polyamines, extracellular Zn 2ϩ , ifenprodil, and alternative splicing, all appear to alter NMDA receptor function by shifting the pKa of the proton sensor (Traynelis et al, 1995;Mott et al, 1998;Choi and Lipton, 1999;Low et al, 2000), our findings have widespread implications for NMDA receptor function.…”

Section: Discussionmentioning

confidence: 99%

“…Allosteric regulators such as polyamines acting at NR1/NR2B NMDA receptors (Traynelis et al, 1995), ifenprodil acting at NR1/NR2B (Mott et al, 1998), and zinc acting at NR1/NR2A (Choi and Lipton, 1999;Low et al, 2000;Paoletti et al, 2000 have been proposed to exert their actions through modification of tonic inhibition by physiological pH. Thus, proton-sensitive gating could be a downstream feature of receptor activation that is regulated by exogenous molecules and therapeutic agents.…”

Section: Introductionmentioning

confidence: 99%

Protons Trap NR1/NR2B NMDA Receptors in a Nonconducting State

Banke¹,

Dravid²,

Traynelis³

2005

J. Neurosci.

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NMDA receptors are highly expressed in the CNS and are involved in excitatory synaptic transmission, as well as synaptic plasticity. Given that overstimulation of NMDA receptors can cause cell death, it is not surprising that these channels are under tight control by a series of inhibitory extracellular ions, including zinc, magnesium, and H ϩ . We studied the inhibition by extracellular protons of recombinant NMDA receptor NR1/NR2B single-channel and macroscopic responses in transiently transfected human embryonic kidney HEK 293 cells using patch-clamp techniques. We report that proton inhibition proceeds identically in the absence or presence of agonist, which rules out the possibility that protonation inhibits receptors by altering coagonist binding. The response of macroscopic currents in excised patches to rapid jumps in pH was used to estimate the microscopic association and dissociation rates for protons, which were 1.4 ϫ 10 9 M Ϫ1 sec Ϫ1 and 110 -196 sec Ϫ1 , respectively (K d corresponds to pH 7.2). Protons reduce the open probability without altering the time course of desensitization or deactivation. Protons appear to slow at least one time constant describing the intra-activation shut-time histogram and modestly reduce channel open time, which we interpret to reflect a reduction in the overall channel activation rate and possible proton-induced termination of openings. This is consistent with a modest proton-dependent slowing of the macroscopic response rise time. From these data, we propose a physical model of proton inhibition that can describe macroscopic and single-channel properties of NMDA receptor function over a range of pH values.

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“…Our previous study (18) as well as more recent work (22) suggested that there may be structural and͞or functional links between inhibition of the NMDA receptor by extracellular protons and Zn 2ϩ . Here, we have explored the nature of the coupling between proton and Zn 2ϩ inhibition and show that there is a unique interaction between Zn 2ϩ and proton inhibition for NR2A-but not NR2B-containing receptors.…”

mentioning

confidence: 88%

“…Here, we have explored the nature of the coupling between proton and Zn 2ϩ inhibition and show that there is a unique interaction between Zn 2ϩ and proton inhibition for NR2A-but not NR2B-containing receptors. We have also used site-directed mutagenesis to systematically probe for coordinate structural determinants of proton and Zn 2ϩ inhibition in the N-terminal leucine͞isoleucine͞valine binding protein (LIVBP)-like domain of NR2A subunit (22,23). This region is critical for the rapidly reversible form of redox modulation (24), which has been suggested to reflect chelation of contaminant Zn 2ϩ (17).…”

mentioning

confidence: 99%

Molecular determinants of coordinated proton and zinc inhibition of N- methyl- d -aspartate NR1/NR2A receptors

Low

Zheng

Lyuboslavsky

et al. 2000

Proc. Natl. Acad. Sci. U.S.A.

157

206

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Modulation of the N-methyl-D-aspartate (NMDA)-selective glutamate receptors by extracellular protons and Zn 2؉ may play important roles during ischemia in the brain and during seizures. Recombinant NR1͞ NR2A receptors exhibit a much higher apparent affinity for voltageindependent Zn 2؉ inhibition than receptors with other subunit combinations. Here, we show that the mechanism of this apparent high-affinity, voltage-independent Zn 2؉ inhibition for NR2A-containing receptors results from the enhancement of proton inhibition. We also show that the N-terminal leucine͞isoleucine͞valine binding protein (LIVBP)-like domain of the NR2A subunit contains critical determinants of the apparent high-affinity, voltage-independent Zn 2؉ inhibition. Mutations H42A, H44G, or H128A greatly increase the Zn 2؉ IC50 (by up to Ϸ700-fold) with no effect on the potencies of glutamate and glycine or on voltage-dependent block by Mg 2؉ . Furthermore, the amino acid residue substitution H128A, which mediates the largest effect on the apparent high-affinity Zn 2؉ inhibition among all histidine substitutions we tested, is also critical to the pH-dependency of Zn 2؉ inhibition. Our data revealed a unique interaction between two important extracellular modulators of NMDA receptors.G lutamate receptors are ligand-gated ion channels that mediate excitatory synaptic transmission in the central nervous system. Although overactivation of the N-methyl-D-aspartate (NMDA)-selective glutamate receptors can trigger neurodegeneration in neuropathological conditions such as stroke, NMDA receptor function is regulated under normal conditions by several extracellular ions (Mg 2ϩ , H ϩ , and Zn 2ϩ ), which exert strong tonic inhibition in a subunit selective manner (1). Inhibition by extracellular protons is particularly relevant for the neuropathological consequences of occlusive stroke because acidification of the extracellular environment during ischemia has been hypothesized to inhibit NMDA receptor overactivation by extrasynaptic glutamate that accumulates following metabolic failure (2, 3). The proton inhibition of NMDA receptors could delay their contribution to subsequent neuronal death until the pH gradients are restored, and this delay may provide a therapeutic window for postinsult treatment with NMDA receptor antagonists. Acidification of the extracellular space during electrographic seizure may also contribute to seizure termination through inhibition of NMDA receptor function (4, 5). Because of these potentially important aspects of the pH sensitivity of the NMDA receptor, we have sought to understand the structural basis by which the receptor might control its regulation by extracellular protons.In the central nervous system, the extracellular Zn 2ϩ concentration has been shown to vary under normal brain function as well as neuropathological conditions (6, 7). In addition, there are large amounts of chelatable Zn 2ϩ in the glutamatergic terminals of hippocampus (8-10), which are released in a Ca 2ϩ -dependent manner (for review, see ref. 6). ...

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Identification and Mechanism of Action of Two Histidine Residues Underlying High-Affinity Zn2+ Inhibition of the NMDA Receptor

Cited by 167 publications

References 44 publications

Structural role of zinc ions bound to postsynaptic densities

Structural role of zinc ions bound to postsynaptic densities

Protons Trap NR1/NR2B NMDA Receptors in a Nonconducting State

Molecular determinants of coordinated proton and zinc inhibition of N- methyl- d -aspartate NR1/NR2A receptors

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