In silico mutational analyses reveal different ligand-binding abilities of double pockets of medaka fish taste receptor type 1 essential for efficient taste recognition.

Taste receptors are important sensors for the detection of nutrient concentrations in animals. Tastes are recognized by interactions between chemical substances and taste receptors. Recently, the high-resolution X-ray crystal structure of the extracellular ligand-binding domains (LBDs) of medaka fish (Oryzias latipes) taste receptor type 1 (T1r) complexed with ligands (amino acids) was determined. Medaka fish T1r is a heterodimer composed of two different LBDs, T1r2aLBD and T1r3LBD. In this study, we performed all-atom molecular dynamics (MD) simulations on this heterodimer (T1r2aLBD-T1r3LBD) to address mutational effects on key residues near each ligand-binding pocket in recognizing one of the ligands (L-Gln). For T1r2aLBD, Ser165 is important in ligand recognition due to its direct hydrogen bonding with the ligand. After mutating Ser165 to Ile or Ala, the direct hydrogen bonds between the ligand and the binding pocket were weakened, which destabilized the ligand-binding form of T1r2aLBD. For T1r3LBD, Ser300 is important in ligand recognition. The water-mediated hydrogen bond with the side-chain hydroxyl group of Ser300 is a single interaction that maintains the ligand-binding form of T1r3LBD. After mutating Ser300 to Glu or Ala, both mutant systems almost maintained their ligand-binding form. As a mechanism for maintaining the binding form of T1r3LBD, alternative hydrogen bonds were formed as direct interactions instead of the indirect water-mediated interactions found in the wild-type system, which stabilized the binding form of T1r3LBD. Judging from our in silico mutational analyses, T1r2aLBD was structurally destabilized by the amino acid mutations. Therefore, it might be required that the ligand-binding pocket of T1r2aLBD is composed of a set of specific residues to maintain its ligand-binding form. On the contrary, T1r3LBD was robust enough to withstand the amino acid mutations. These different ligand-binding abilities of both LBDs provide multiple binding modes, which might be helpful for discriminating various taste substances or detecting concentrations of nutrients efficiently.