How Can Mutations Thermostabilize G-Protein-Coupled Receptors?

Structures of over 30 different G-protein-coupled receptors (GPCRs) have advanced our understanding of cell signaling and have provided a foundation for structure-guided drug design. This exciting progress has required the development of three complementary methods to facilitate GPCR crystallization, one of which is the thermostabilization of receptors by systematic mutagenesis. However, the reason why a particular mutation, or combination of mutations, stabilizes the receptor is not always evident from a static crystal structure. Molecular dynamics (MD) simulations have been used to identify and estimate the energetic factors that affect thermostability through comparing the dynamics of the thermostabilized receptors with structure-based models of the wild-type receptor. The data indicate that receptors are stabilized through a combination of factors, including an increase in receptor rigidity, a decrease in collective motion, reduced stress at specific residues, and the presence of ordered water molecules. Predicting thermostabilizing mutations computationally represents a major challenge for the field.

TrendsGPCRs are major drug targets and are involved in mediating multiple signal transduction pathways in the cell.One of the successful strategies that lead to solving many GPCR crystal structures in different conformations is thermostabilization of the receptor by systematic mutagenesis.The antagonist-bound thermostable mutants in their inactive state show receptor pharmacologies similar to those of their respective wild-type receptors.The agonist-bound thermostable mutants are more dynamic and show conformational variations in the extracellular half of the receptor depending on the agonist-bound than in the intracellular half.MD simulations show that thermostable mutants are less flexible than their wild-type counterparts.Most of the thermostable mutant GPCRs gain their stability from enthalpy stemming from improved interhelical packing compared to wild-type receptors.