Psychedelics promote plasticity by directly binding to BDNF receptor TrkB.

Psychedelics produce fast and persistent antidepressant effects and induce neuroplasticity resembling the effects of clinically approved antidepressants. We recently reported that pharmacologically diverse antidepressants, including fluoxetine and ketamine, act by binding to TrkB, the receptor for BDNF. Here we show that lysergic acid diethylamide (LSD) and psilocin directly bind to TrkB with affinities 1,000-fold higher than those for other antidepressants, and that psychedelics and antidepressants bind to distinct but partially overlapping sites within the transmembrane domain of TrkB dimers. The effects of psychedelics on neurotrophic signaling, plasticity and antidepressant-like behavior in mice depend on TrkB binding and promotion of endogenous BDNF signaling but are independent of serotonin 2A receptor (5-HT2A) activation, whereas LSD-induced head twitching is dependent on 5-HT2A and independent of TrkB binding. Our data confirm TrkB as a common primary target for antidepressants and suggest that high-affinity TrkB positive allosteric modulators lacking 5-HT2A activity may retain the antidepressant potential of psychedelics without hallucinogenic effects.

NanoLuc Luciferase as a Fluorogen-Activating Protein for GFP Chromophore Based Fluorogens.

In this work, we showed that the well-known NanoLuc luciferase can act as a fluorogen activating protein for various arylidene-imidazolones structurally similar to the Kaede protein chromophore. We showed that such compounds can be used as fluorescent sensors for this protein and can also be used in pairs with it in fluorescent microscopy as a genetically encoded tag.

Investigation of lipid/protein interactions in trifluoroethanol-water mixtures proposes the strategy for the refolding of helical transmembrane domains.

Membrane proteins are one of the keystone objects in molecular biology, but their structural studies often require an extensive search for an appropriate membrane-like environment and an efficient refolding protocol for a recombinant protein. Isotropic bicelles are a convenient membrane mimetic used in structural studies of membrane proteins. Helical membrane domains are often transferred into bicelles from trifluoroethanol-water mixtures. However, the protocols for such a refolding are empirical and the process itself is still not understood in detail. In search of the optimal refolding approaches for helical membrane proteins, we studied here how membrane proteins, lipids, and detergents interact with each other at various trifluoroethanol-water ratios. Using high-resolution NMR spectroscopy and dynamic light scattering, we determined the key states of the listed compounds in the trifluoroethanol/water mixture, found the factors that could be critical for the efficiency of refolding, and proposed several most optimal protocols. These protocols were developed on the transmembrane domain of neurotrophin receptor TrkA and tested on two model helical membrane domains-transmembrane of Toll-like receptor TLR9 and voltage-sensing domain of a potassium channel KvAP.

Indane Based Molecular Motors: UV-Switching Increases Number of Isomers.

We describe azophenylindane based molecular motors (aphin-switches) which have two different rotamers of trans-configuration and four different rotamers of cis-configuration. The behaviors of these motors were investigated both experimentally and computationally. The conversion of aphin-switch does not yield single isomer but a mixture of these. Although the trans to cis conversion leads to the increase of the system entropy some of the cis-rotamers can directly convert to each other while others should convert via trans-configuration. The motion of aphin-switches resembles the work of a mixing machine with indane group serving as a base and phenol group serving as a beater. The aphin-switches presented herein may provide a basis for promising applications in advanced biological systems or particularly in cases where on demand disordering of molecular packing has value, such as lipid bilayers.

Intrinsically disordered regions couple the ligand binding and kinase activation of Trk neurotrophin receptors.

Receptor tyrosine kinases (RTKs) are key players in development and several diseases. Understanding the molecular mechanism of RTK activation by its ligand could lead to the design of new RTK inhibitors. How the extracellular domain is coupled to the intracellular kinase domain is a matter of debate. Ligand-induced dimerization and ligand-induced conformational change of pre-formed dimers are two of the most proposed models. Recently we proposed that TrkA, the RTK for nerve growth factor (NGF), is activated by rotation of the transmembrane domain (TMD) pre-formed dimers upon NGF binding. However, one of the unsolved issues is how the ligand binding is conformationally coupled to the TMD rotation if unstructured extracellular juxtamembrane (eJTM) regions separate them. Here we use nuclear magnetic resonance in bicelles and functional studies to demonstrate that eJTM regions from the Trk family are intrinsically disordered and couple the ligand-binding domains and TMDs possibly via the interaction with NGF.

Oligomerization analysis as a tool to elucidate the mechanism of EBV latent membrane protein 1 inhibition by pentamidine.

Latent membrane protein 1 (LMP1) is a gene product of the Epstein-Barr virus (EBV), a widely spread virus present in 90-95% of the world's population. EBV can lead to several malignancies, in which LMP1 was shown to play a key role. LMP1 is active only in the oligomeric form and its fifth transmembrane domain (TMD-5) is critical for the oligomerization, with D150 identified as a key residue for LMP1 activation. Here we propose an NMR-based approach to treat the complex oligomerization equilibria with slow conformational exchange. Using this method we investigate the TMD-5 in DPC micelles. We show that the pKa of D150 equals 7.4. Uncharged form of TMD-5 associates into dimers and trimers, deprotonation of D150 induces the high-order oligomerization of the protein and enhances dramatically its trimerization. Pentamidine interacts mainly with the charged TMD-5, destroying the oligomers and stabilizing the monomer and trimer. Using computer simulations we investigate the structural basis of TMD-5/pentamidine interaction. Our data suggest that D150 is likely charged in the full-length LMP1 under native conditions.

Phase Transitions in Small Isotropic Bicelles.

Isotropic phospholipid bicelles are one of the most prospective membrane mimetics for the structural studies of membrane proteins in solution. Recent works provided an almost full set of data regarding the properties of isotropic bicelles; however, one major aspect of their behavior is still under consideration: the possible mixing between the lipid and detergent in the bilayer area. This problem may be resolved by studying the lipid phase transitions in bicelle particles. In the present work, we investigate two effects: phase transitions of bilayer lipids and temperature-induced growth of isotropic bicelles using the NMR spectroscopy. We propose an approach to study the phase transitions in isotropic bicelles based on the properties of 31P NMR spectra of bilayer-forming lipids. We show that phase transitions in small bicelles are "fractional", particles with the liquid-crystalline and gel bilayers coexist in solution at certain temperatures. We study the effects of lipid fatty chain type and demonstrate that the behavior of various lipids in bilayers is reproduced in the isotropic bicelles. We show that the temperature-induced growth of isotropic bicelles is not related directly to the phase transition but is the result of the reversible fusion of bicelle particles. In accordance with our data, rim detergents also have an impact on phase transitions: detergents that resist the temperature-induced growth provide the narrowest and most expressed transitions at higher temperatures. We demonstrate clearly that phase transitions take place even in the smallest bicelles that are applicable for structural studies of membrane proteins by solution NMR spectroscopy. This last finding, together with other data draws a thick line under the long-lasting argument about the relevance of small isotropic bicelles. We show with certainty that the small bicelles can reproduce the most fundamental property of lipid membranes: the ability to undergo phase transition.