Summary: Computer simulations are giving way to more complex and accurate studies of biological membranes by molecular dynamics (MD) simulations. The analysis of MD trajectories comprises the biophysical characterization of membrane properties or the study of protein–lipid interactions and dynamics. However, there is a lack of automated tools to analyse MD simulations of complex membrane or membrane-protein systems. Here we present MEMBPLUGIN, a plugin for the Visual Molecular Dynamics package that provides algorithms to measure a host of essential biophysical properties in simulated membranes. MEMBPLUGIN features are accessible both through a user-friendly graphical interface and as command-line procedures to be invoked in analysis scripts. Availability and implementation: MEMBPLUGIN is a VMD extension written in Tcl. Multi-platform source code, documentation and tutorials are freely available at http://membplugin.sourceforge.net. Contact:toni.giorgino@isib.cnr.it or jana.selent@upf.edu Supplementary information: Supplementary data are available at Bioinformatics online.

G protein-coupled receptors modulate a plethora of physiological processes and mediate the effects of one-third of FDA-approved drugs. Depending on which ligand activates a receptor, it can engage different intracellular transducers. This ‘biased signaling’ paradigm requires that we now characterize physiological signaling not just by receptors but by ligand-receptor pairs. Ligands eliciting biased signaling may constitute better drugs with higher efficacy and fewer adverse effects. However, ligand bias is very complex, making reproducibility and description challenging. Here, we provide guidelines and terminology for any scientists to design and report ligand bias experiments. The guidelines will aid consistency and clarity, as the basic receptor research and drug discovery communities continue to advance our understanding and exploitation of ligand bias. Scientific insight, biosensors, and analytical methods are still evolving and should benefit from and contribute to the implementation of the guidelines, together improving translation from in vitro to disease-relevant in vivo models.

Cholesterol is a key component of cell membranes with a proven modulatory role on the function and ligand-binding properties of G-protein-coupled receptors (GPCRs). Crystal structures of prototypical GPCRs such as the adenosine A_2A receptor (A_2AR) have confirmed that cholesterol finds stable binding sites at the receptor surface suggesting an allosteric role of this lipid. Here we combine experimental and computational approaches to show that cholesterol can spontaneously enter the A_2AR-binding pocket from the membrane milieu using the same portal gate previously suggested for opsin ligands. We confirm the presence of cholesterol inside the receptor by chemical modification of the A_2AR interior in a biotinylation assay. Overall, we show that cholesterol’s impact on A_2AR-binding affinity goes beyond pure allosteric modulation and unveils a new interaction mode between cholesterol and the A_2AR that could potentially apply to other GPCRs.

Membrane levels of docosahexaenoic acid (DHA), an essential omega-3 polyunsaturated fatty acid (ω-3 PUFA), are decreased in common neuropsychiatric disorders. DHA modulates key cell membrane properties like fluidity, thereby affecting the behaviour of transmembrane proteins like G protein-coupled receptors (GPCRs). These receptors, which have special relevance for major neuropsychiatric disorders have recently been shown to form dimers or higher order oligomers and evidence suggests that DHA levels affect GPCR function by modulating oligomerisation. In this study, we assessed the effect of membrane DHA content on the formation of a class of protein complexes with particular relevance for brain disease: adenosine A_2A and dopamine D₂ receptor oligomers. Using extensive multiscale computer modelling, we find a marked propensity of DHA for interaction with both A_2A and D₂ receptors, which leads to an increased rate of receptor oligomerisation. Bioluminescence resonance energy transfer (BRET) experiments performed on living cells suggest that this DHA effect on the oligomerisation of A_2A and D₂ receptors is purely kinetic. This work reveals for the first time that membrane ω-3 PUFAs play a key role in GPCR oligomerisation kinetics, which may have important implications for neuropsychiatric conditions like schizophrenia or Parkinson’s disease.