Ion-translocating rhodopsins, especially channelrhodopsins (ChRs), have attracted broad attention as a powerful tool to modulate the membrane potential of cells with light (optogenetics). Because of ...recent biophysical, spectroscopic, and computational studies, including the structural determination of cation and anion ChRs, our understanding of the molecular mechanism underlying light-gated ion conduction has been greatly advanced. In this chapter, I first describe the background of rhodopsin family proteins including ChR, and how the optogenetics technology has been established from the discovery of first ChR in 2002. I later introduce the recent findings of the structure-function relationship of ChR by comparing the crystal structures of cation and anion ChRs. I further discuss the future goal in the fields of ChR research and optogenetic tool development.
Cannabis elicits its mood-enhancing and analgesic effects through the cannabinoid receptor 1 (CB1), a G protein-coupled receptor (GPCR) that signals primarily through the adenylyl cyclase-inhibiting ...heterotrimeric G protein Gi. Activation of CB1-Gi signaling pathways holds potential for treating a number of neurological disorders and is thus crucial to understand the mechanism of Gi activation by CB1. Here, we present the structure of the CB1-Gi signaling complex bound to the highly potent agonist MDMB-Fubinaca (FUB), a recently emerged illicit synthetic cannabinoid infused in street drugs that have been associated with numerous overdoses and fatalities. The structure illustrates how FUB stabilizes the receptor in an active state to facilitate nucleotide exchange in Gi. The results compose the structural framework to explain CB1 activation by different classes of ligands and provide insights into the G protein coupling and selectivity mechanisms adopted by the receptor.
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•3 Å cryo-EM structure of the CB1-Gi complex bound to potent agonist MDMB-Fubinaca•MDMB-Fubinaca locks “toggle switch” residues F2003.36/W3566.48 in active conformation•Quantum mechanics calculations reveal the mechanism for the high affinity of Fubinaca•Molecular dynamic simulations reveal a path for ligand entry between TM1 and TM7
Looking at how a toxic, synthetic ligand locks cannabinoid receptor 1 into a signaling conformation points to ways to understand and modulate receptor activity.
Perceptual experiences may arise from neuronal activity patterns in mammalian neocortex. We probed mouse neocortex during visual discrimination using a red-shifted channelrhodopsin (ChRmine, ...discovered through structure-guided genome mining) alongside multiplexed multiphoton-holography (MultiSLM), achieving control of individually specified neurons spanning large cortical volumes with millisecond precision. Stimulating a critical number of stimulus-orientation-selective neurons drove widespread recruitment of functionally related neurons, a process enhanced by (but not requiring) orientation-discrimination task learning. Optogenetic targeting of orientation-selective ensembles elicited correct behavioral discrimination. Cortical layer-specific dynamics were apparent, as emergent neuronal activity asymmetrically propagated from layer 2/3 to layer 5, and smaller layer 5 ensembles were as effective as larger layer 2/3 ensembles in eliciting orientation discrimination behavior. Population dynamics emerging after optogenetic stimulation both correctly predicted behavior and resembled natural internal representations of visual stimuli at cellular resolution over volumes of cortex.
Proton-dependent oligopeptide transporters (POTs) are major facilitator superfamily (MFS) proteins that mediate the uptake of peptides and peptide-like molecules, using the inwardly directed H ⁺ ...gradient across the membrane. The human POT family transporter peptide transporter 1 is present in the brush border membrane of the small intestine and is involved in the uptake of nutrient peptides and drug molecules such as β-lactam antibiotics. Although previous studies have provided insight into the overall structure of the POT family transporters, the question of how transport is coupled to both peptide and H ⁺ binding remains unanswered. Here we report the high-resolution crystal structures of a bacterial POT family transporter, including its complex with a dipeptide analog, alafosfalin. These structures revealed the key mechanistic and functional roles for a conserved glutamate residue (Glu310) in the peptide binding site. Integrated structural, biochemical, and computational analyses suggested a mechanism for H ⁺-coupled peptide symport in which protonated Glu310 first binds the carboxyl group of the peptide substrate. The deprotonation of Glu310 in the inward open state triggers the release of the bound peptide toward the intracellular space and salt bridge formation between Glu310 and Arg43 to induce the state transition to the occluded conformation.
Krokinobacter eikastus rhodopsin 2 (KR2) is the first light-driven Na(+) pump discovered, and is viewed as a potential next-generation optogenetics tool. Since the positively charged Schiff base ...proton, located within the ion-conducting pathway of all light-driven ion pumps, was thought to prohibit the transport of a non-proton cation, the discovery of KR2 raised the question of how it achieves Na(+) transport. Here we present crystal structures of KR2 under neutral and acidic conditions, which represent the resting and M-like intermediate states, respectively. Structural and spectroscopic analyses revealed the gating mechanism, whereby the flipping of Asp116 sequesters the Schiff base proton from the conducting pathway to facilitate Na(+) transport. Together with the structure-based engineering of the first light-driven K(+) pumps, electrophysiological assays in mammalian neurons and behavioural assays in a nematode, our studies reveal the molecular basis for light-driven non-proton cation pumps and thus provide a framework that may advance the development of next-generation optogenetics.
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Dostopno za:
DOBA, IJS, IZUM, KILJ, KISLJ, NUK, PILJ, PNG, SAZU, SBMB, SIK, UILJ, UKNU, UL, UM, UPUK
Neurotensin receptor 1 (NTSR1) is a G-protein-coupled receptor (GPCR) that engages multiple subtypes of G protein, and is involved in the regulation of blood pressure, body temperature, weight and ...the response to pain. Here we present structures of human NTSR1 in complex with the agonist JMV449 and the heterotrimeric Gi1 protein, at a resolution of 3 Å. We identify two conformations: a canonical-state complex that is similar to recently reported GPCR-Gi/o complexes (in which the nucleotide-binding pocket adopts more flexible conformations that may facilitate nucleotide exchange), and a non-canonical state in which the G protein is rotated by about 45 degrees relative to the receptor and exhibits a more rigid nucleotide-binding pocket. In the non-canonical state, NTSR1 exhibits features of both active and inactive conformations, which suggests that the structure may represent an intermediate form along the activation pathway of G proteins. This structural information, complemented by molecular dynamics simulations and functional studies, provides insights into the complex process of G-protein activation.
Pulsar magnetospheres have strong magnetic fields and large amounts of plasma. The structures of these magnetospheres are studied using force-free electrodynamics. To understand pulsar ...magnetospheres, discussions must include their outer region. However, force-free electrodynamics is limited in it does not handle dissipation. Therefore, a resistive pulsar magnetic field model is needed. To break the ideal magnetohydrodynamic (MHD) condition , Ohm's law is used. This work introduces resistivity depending upon the distance from the star and obtain a self-consistent steady state by time integration. Poloidal current circuits form in the magnetosphere while the toroidal magnetic field region expands beyond the light cylinder and the Poynting flux radiation appears. High electric resistivity causes a large space scale poloidal current circuit and the magnetosphere radiates a larger Poynting flux than the linear increase outside of the light cylinder radius. The formed poloidal-current circuit has width, which grows with the electric conductivity. This result contributes to a more concrete dissipative pulsar magnetosphere model.
Channelrhodopsins (ChRs) are light-gated cation channels derived from algae that have shown experimental utility in optogenetics; for example, neurons expressing ChRs can be optically controlled with ...high temporal precision within systems as complex as freely moving mammals. Although ChRs have been broadly applied to neuroscience research, little is known about the molecular mechanisms by which these unusual and powerful proteins operate. Here we present the crystal structure of a ChR (a C1C2 chimaera between ChR1 and ChR2 from Chlamydomonas reinhardtii) at 2.3 Å resolution. The structure reveals the essential molecular architecture of ChRs, including the retinal-binding pocket and cation conduction pathway. This integration of structural and electrophysiological analyses provides insight into the molecular basis for the remarkable function of ChRs, and paves the way for the precise and principled design of ChR variants with novel properties.
The authors used a longitudinal impact evaluation with quasi-experimental methods to provide evidence on economic and production impact of a farmer field school (FFS) project in East Africa. FFSs ...were shown to have positive impact on production and income among women, low-literacy, and medium land size farmers. Participation in FFS increased income by 61%. Participation in FFS improved agricultural income and crop productivity overall. This implies that farmer field schools are a useful approach to increase production and income of small-scale farmers in East Africa, and that the approach can be used to target women and producers with limited literacy.
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