Conformational plasticity of ligand-bound and ternary GPCR complexes studied by F NMR of the β-adrenergic receptor.

G-protein-coupled receptors (GPCRs) are allosteric signaling proteins that transmit an extracellular stimulus across the cell membrane. Using F NMR and site-specific labelling, we investigate the response of the cytoplasmic region of transmembrane helices 6 and 7 of the β-adrenergic receptor to agonist stimulation and coupling to a G-protein-mimetic nanobody. Agonist binding shows the receptor in equilibrium between two inactive states and a pre-active form, increasingly populated with higher ligand efficacy.

The role of NMR spectroscopy in mapping the conformational landscape of GPCRs.

Over recent years, nuclear magnetic resonance (NMR) spectroscopy has developed into a powerful mechanistic tool for the investigation of G protein-coupled receptors (GPCRs). NMR provides insights which underpin the dynamic nature of these important receptors and reveals experimental evidence for a complex conformational energy landscape that is explored during receptor activation resulting in signalling. NMR studies have highlighted both the dynamic properties of different receptor states as well as the exchange pathways and intermediates formed during activation, extending the static view of GPCRs obtained from other techniques.

Evolthon: A community endeavor to evolve lab evolution.

In experimental evolution, scientists evolve organisms in the lab, typically by challenging them to new environmental conditions. How best to evolve a desired trait? Should the challenge be applied abruptly, gradually, periodically, sporadically? Should one apply chemical mutagenesis, and do strains with high innate mutation rate evolve faster? What are ideal population sizes of evolving populations? There are endless strategies, beyond those that can be exposed by individual labs. We therefore arranged a community challenge, Evolthon, in which students and scientists from different labs were asked to evolve Escherichia coli or Saccharomyces cerevisiae for an abiotic stress-low temperature.

Insight into partial agonism by observing multiple equilibria for ligand-bound and G-mimetic nanobody-bound β-adrenergic receptor.

A complex conformational energy landscape determines G-protein-coupled receptor (GPCR) signalling via intracellular binding partners (IBPs), e.g., G and β-arrestin.

An Adaptable Phospholipid Membrane Mimetic System for Solution NMR Studies of Membrane Proteins.

Based on the saposin-A (SapA) scaffold protein, we demonstrate the suitability of a size-adaptable phospholipid membrane-mimetic system for solution NMR studies of membrane proteins (MPs) under close-to-native conditions. The Salipro nanoparticle size can be tuned over a wide pH range by adjusting the saposin-to-lipid stoichiometry, enabling maintenance of sufficiently high amounts of phospholipid in the Salipro nanoparticle to mimic a realistic membrane environment while controlling the overall size to enable solution NMR for a range of MPs. Three representative MPs, including one G-protein-coupled receptor, were successfully incorporated into SapA-dimyristoylphosphatidylcholine nanoparticles and studied by solution NMR spectroscopy.