Photoswitchable Amino Acids For Control Of NMDA Receptor Activity

Nerve cells communicate with each other by releasing chemicals, known as neurotransmitters, from one cell to the next. Once released, these neurotransmitters bind to specific docking stations, called receptors, which are located on the surface of the neighboring cell.

Due to changes in neurotransmitter release or the receptor number, the connections between neurons can either strengthen or weaken over time. This process, called synaptic plasticity, forms the basis of learning and memory. One of the key players in synaptic plasticity are N-methyl-D-aspartate (NMDA) receptors, and if these receptors are faulty, it can cause disorders such as schizophrenia or epilepsy.

NMDAs are a large family of receptors that have many receptor subtypes, each with specific properties. Every subtype is composed of four varying subunits. It is still unclear how these different receptor subtypes contribute to synaptic plasticity and new methods are needed to resolve this puzzle.

Light Switch Molecule

An emerging strategy to study brain receptors, known as optogenetics, is to engineer them so that they can be controlled with light. One approach to provide light-sensitivity uses molecules that act as ‘light switches’.

These switches change their shape when exposed to specific colors of light and this way, turn a receptor on or off. However, commonly used light switches are often very large, meaning that they can only be introduced at specific sites in a receptor, and have limited ability to change the shape of a receptor.

Proposed mechanism for photomodulation of the gating and permeation properties of NMDARs carrying PSAAs.

Proposed mechanism for photomodulation of the gating and permeation properties of NMDARs carrying PSAAs.
(a) Toggling of the PSAA azobenzene moiety between the trans- and cis-isomers in dependence of blue or UV light. For simplicity, the two isomers are replaced by symbols as indicated.
(b) Schematic representation of the mechanism underlying the photomodulation at the ABD level. For clarity, only one dimer is represented and the NTD region is omitted. PSAA in its trans-configuration (dark and blue condition), when placed at the GluN1 ABD clamshell hinge (P532 position), intrudes into a hydrophobic pocket located at the GluN2A ABD hinge region (Furukawa et al., 2005). This interaction stabilizes the GluN1 ABD in a closed-cleft conformation (i.e. active state) as well as ABD GluN1/GluN2 heterodimer interface, resulting in active (i.e. open) channels (Furukawa et al., 2005; Gielen et al., 2008). Flipping the GluN1-azobenzene moiety to the cis-configuration by UV illumination destabilizes this active conformation, favoring GluN1 clamshell opening, which in turn accelerates glycine dissociation. This effect is accompanied by a weakening of the ABD dimer interface resulting in a 50% drop of channel Po (UV-inhibited state).
(c) Schematic representation of the mechanism underlying photomodulation in the pore domain. The TMD pore region is viewed from the top. PSAA is placed at the GluN1-Y647 site, which is part of a highly conserved hydrophobic cluster that sits at the back of the channel lumen and connects the M3 helix bundle (channel gate, inner ring) to the peripheral M1/M4 helix outer ring. PSAA in its extended trans-configuration prohibits adequate channel opening by preventing full expansion of inner M3 ring towards the periphery. The channel is thus locked in a constrained ‘open’ state, highly unstable (low Po) and with low conductance. Upon UV, however, activity can be drastically enhanced, an effect that originates from the relief of the steric hindrance allowing the inner M3 ring to accomplish its full movement towards the outer ring (UV-activated state). This motion generates an increase in ion channel Po and conductance. Thus, channel opening is energetically more favorable in the more compact cis-configuration than the rigid and elongated trans-configuration of the PSAA.

Viktoria Klippenstein, of the Institut de Biologie de l’École Normale Supérieure, and colleagues have now generated a small light switch molecule with the size of a single amino acid side-chain that, in theory, could replace any of the usual amino acids in the NMDA receptor.

Different locations for the light switch were tested to identify those that changed the activity of the receptor. When the receptors were stimulated with light, the light switch changed its shape, which in turn influenced the shape of the receptor.

This meant that, depending on which amino acid in the receptor had been replaced with the light switch, light could be used to control the receptor activity in different ways.

This new approach of using integrated light switches allows NMDA receptors to be controlled in a fast and reversible manner using something as simple as a beam of light. Further research will use the tool set of light-controllable receptors to study how the different NMDA receptor subtypes affect synaptic plasticity in the normal and diseased brain.

Funding for the research was provided by Agence Nationale de la Recherche, Centre National de la Recherche Scientifique, Institut National de la Santé et de la Recherche Médicale, National Institutes of Health, European Research Council, Fondation pour la Recherche Médicale, and Deutsche Forschungsgemeinschaft.

Viktoria Klippenstein, Christian Hoppmann, Shixin Ye, Lei Wang, Pierre Paoletti
Optocontrol of glutamate receptor activity by single side-chain photoisomerization
eLife 2017;6:e25808

© 2017, Klippenstein et al. Republished under the terms of the Creative Commons Attribution License.
Top Image: Michael Northrop, Wellcome Images