Biology subjects

Koers, E. J.

Publications and source records attributed to Koers, E. J..

4 recordsLinked to original sources

bioRxiv2021

Individual cells traffic the Vasopressin 2 Receptor to their cell surface with different success

G protein coupled receptors (GPCRs) translate the actions of hormones into intracellular signalling events. Mutations in GPCRs can prevent their correct expression and trafficking to the cell surface and cause disease. We use single cell measurements in HEK293 cells to show that the balance between endoplasmic reticulum (ER) and cell surface localisation of the Vasopressin 2 receptor (V2R) varies significantly from cell to cell. We find that mutations in the V2R affect the proportion of cells able to send this GPCR to the cell surface but do not prevent all cells in the population from correctly trafficking the mutant receptors. These findings reveal that the ability of cells to correctly traffic V2R to the cell surface depends not only on the expressed V2R mutant but also on the individual cell environment. Significance statementMissense mutations in the Vasopressin 2 Receptor (V2R) cause Nephrogenic Diabetes Insipidus. Some of these mutations prevent correct expression and trafficking of V2R to the cell surface resulting in a loss-of-function. We show -using single cell measurements-that the balance between endoplasmic reticulum and cell surface localisation of the V2R varies significantly from cell to cell, independent from its expression level. Mutations affect the proportion of cells able to send V2R to the cell surface but do not prevent all cells in the population from correctly trafficking the mutant receptors. Hence, the ability of cells to correctly traffic V2R to the cell surface depends not only on expressed V2R mutant but also on the cell environment.

molecular biology↗

bioRxiv2021

Direct observation of Hsp90-induced compaction in a protein chain

The chaperone Hsp90 is well known to undergo important conformational changes, which depend on nucleotide, co-chaperones, substrate interactions and post-translational modifications. Conversely, how the conformations of its unstable and disordered substrates are affected by Hsp90 is difficult to address experimentally, yet central to its function. Here, using optical tweezers and luciferase and glucocorticoid receptor substrates, we find that Hsp90 promotes local contractions in unfolded chains that drive their global compaction down to dimensions of folded states. This compaction has a gradual nature while showing small steps, is stimulated by ATP, and performs mechanical work against counteracting forces that expand the chain dimensions. The Hsp90 interactions suppress the formation of larger-scale folded, misfolded and aggregated structures. The observations support a model in which Hsp90 alters client conformations directly by promoting local intra-chain interactions while suppressing distant ones. We conjecture that chain compaction may be central to how Hsp90 protects unstable kinases and receptor clients, regulates their activity, and how Hsp90 cooperates with Hsp70.

biophysics↗

bioRxiv2021

Prediction of ligand-receptor pharmacological activities using a combined docking and machine learning approach

G protein coupled receptors (GPCRs) are valuable therapeutic targets for many diseases. A central question of GPCR drug discovery is to understand what determines the agonism or antagonism of ligands which bind them. Ligands exert their action via the interactions in the ligand binding pocket. We hypothesised that there is a common set of receptor interactions made by ligands of diverse structures that mediate their action and that among a large dataset of different ligands, the functionally important interactions will be over-represented. We computationally docked ~2700 known {beta}2AR ligands to multiple {beta}2AR structures, generating ca 75,000 docking poses and predicted all atomic interactions between the receptor and the ligand. We used machine learning (ML) techniques to identify specific interactions that correlate with the agonist or antagonist activity of these ligands. The interpretation of ML analysis in human understandable form allowed us to construct an exquisitely detailed structure-activity relationship that identifies small changes to the ligands that invert their activity and thus helps to guide the drug discovery process. This approach can be readily applied to any drug target.

bioinformatics↗

bioRxiv2020

Polypeptide collapse modulation and folding stimulation by GroEL-ES

Unfolded proteins ubiquitously collapse into a compact yet dynamic state1,2. While this compaction is pivotal to protein folding3, aggregation4,5, intrinsic disorder6, and phase separation7, its role in protein quality control mechanisms remains obscure8. Collapse has been characterized mainly for polypeptides that are free in solution, in terms of kinetics, chain expansion, and effect on folding9,10. Yet, theory suggests that the solvent-mediated forces driving collapse can be altered near hydrophobic and charged surfaces, which are observed for many proteins including GroEL-ES11,12. Notably, while GroEL-ES is the archetypal protein-folding chaperone, its folding mechanism remains unresolved13,14. GroEL-ES is proposed to sterically confine polypeptides within its closed chamber15, unfold misfolded states16,17, or promote folding indirectly by suppressing aggregation18,19. Here, using integrated protein manipulation and imaging, we show that GroEL-ES can strengthen the collapse of polypeptide substrates, and hence stimulate folding directly. Strikingly, attractive forces pull substrate chains into the open GroEL cavity -unclosed by GroES-, and hence trigger a gradual compaction and discrete folding transitions, even for slow-folding proteins. This collapse enhancement is strongest in the nucleotide-bound states of GroEL, and is aided by GroES binding to the cavity rim, and by the amphiphilic C-terminal tails at the cavity bottom. Peptides corresponding to these C-termini alone are sufficient to strengthen the collapse. The results show a mechanism that allows folding to be stimulated: by strengthening the collapse, residues are brought together that must contact to fold. The notion that one protein can modulate the collapse of another may be generally important in protein conformation and coacervation control, for systems ranging from the GroEL-ES homologue TRiC/CCT20, to the oncogenic c-Myc/Max complex21, and the nuclear pore transporter transportin22.

biophysics↗