Conformational thermostabilization of the β1-adrenergic receptor in a detergent-resistant form

TLDR: A generic strategy for the isolation of detergent-solubilized thermostable mutants of a GPCR, the β1-adrenergic receptor, and the most stable mutant receptor, βAR-m23, was significantly more stable in a wide range of detergents ideal for crystallization and was preferentially in an antagonist conformation in the absence of ligand.

Abstract: There are ≈350 non-odorant G protein-coupled receptors (GPCRs) encoded by the human genome, many of which are predicted to be potential therapeutic targets, but there are only two structures available to represent the whole of the family. We hypothesized that improving the detergent stability of these receptors and simultaneously locking them into one preferred conformation will greatly improve the chances of crystallization. We developed a generic strategy for the isolation of detergent-solubilized thermostable mutants of a GPCR, the β1-adrenergic receptor. The most stable mutant receptor, βAR-m23, contained six point mutations that led to an apparent Tm 21°C higher than the native protein, and, in the presence of bound antagonist, βAR-m23 was as stable as bovine rhodopsin. In addition, βAR-m23 was significantly more stable in a wide range of detergents ideal for crystallization and was preferentially in an antagonist conformation in the absence of ligand.