Sensitivity of human sweet taste receptor subunits T1R2 and T1R3 to activation by glucose enantiomers

Abstract

We have previously shown that L-glucose, the non-caloric enantiomer of D-glucose, activates the human sweet taste receptor T1R2/T1R3 transiently expressed in HEK293T cells. Here we show that D- and L-glucose can also activate T1R2 and T1R3 expressed without the counterpart monomer. Serine mutation to alanine in residue 147 in the binding site of T1R3 VFT domain, completely abolishes T1R3S147A activation by either L or D-glucose, while T1R2/T1R3S147A responds in the same way as T1R2 expressed without its counterpart. We further show that the original T1R2 reference sequence (NM_152232.1) is less sensitive by almost an order of magnitude than reference sequence at the time this study was performed (NM_152232.4). We find that out of the four differing positions, it is the R317G in the VFT domain of T1R2, that is responsible for this effect in-vitro. It is important both for practical assay sensitivity, and because glycine is found in this position in ~20% of the world population. While the effects of the mutations and of the partial transfections were similar for D and L enantiomers, their dose response curves remained distinct, with L-glucose reaching an early plateau.

T1R3 subunit of the sweet taste receptor is activated by D2O in transmembrane domain-dependent manner

“…2A). Dubovski et al showed that the T1R2 (NM_152232.4) is more sensitive to presence of D-glucose 10 . In order to understand whether these SNPs affect the sensitivity of activation by D 2 O, we compared the dose response curves of HEK293T cells, transfected with either the T1R2 (NM_152232.1) or the more sensitive version (NM_152232.4), along with T1R3.…”

“…Varying concentrations of deuterium Oxide were tested among different transfections; T1R2(NM_152232.1)/T1R3(black), T1R3/T1R3(blue) or T1R2/T1R2(NM_152232.1, pink). (B) Dose-response curves of HEK293T cells transiently transfected with T1R2(NM_152232.1)/T1R3 (black); or T1R2(NM_152232.4)/T1R3 (grey); All data were normalized based on maximum response of the full sweet taste heterodimer receptor 10 T1R2/T1R3 to 0.96M glucose. All tested solutions were done in at least triplicate; each experiment (transfection) was repeated three to five times.…”

“…Activation of the receptor was monitored by IP1 accumulation. 960mM of D--glucose was used here as control for each transfection, based on previous work 4, 10 .…”

“…Residue R317 in the VFT domain of T1R2, was shown to be responsible for this difference. Furthermore, they showed that site--directed mutagenesis of Serine at position 147 in the binding site of T1R3 VFT domain to Alanine, abolished receptor activation by D and L glucose enantiomers 10 .…”

“…Interestingly, T1R3 without T1R2 (possibly functioning as a homodimer) was shown to respond to monosaccharides and disaccharides 10, 39 . This is of physiological relevance, since T1R3 (but not T1R2) expression was shown in the gastrointestinal tract 25 pancreas and liver 9, 31 .…”