Characterization of the Binding Site of Aspartame in the Human Sweet Taste Receptor

Abstract

The sweet taste receptor, a heterodimeric G protein-coupled receptor comprised of T1R2 and T1R3, binds sugars, small molecule sweeteners, and sweet proteins to multiple binding sites. The dipeptide sweetener, aspartame binds in the Venus Flytrap Module (VFTM) of T1R2. We developed homology models of the open and closed forms of human T1R2 and human T1R3 VFTMs and their dimers and then docked aspartame into the closed form of T1R2's VFTM. To test and refine the predictions of our model, we mutated various T1R2 VFTM residues, assayed activity of the mutants and identified 11 critical residues (S40, Y103, D142, S144, S165, S168, Y215, D278, E302, D307, and R383) in and proximal to the binding pocket of the sweet taste receptor that are important for ligand recognition and activity of aspartame. Furthermore, we propose that binding is dependent on 2 water molecules situated in the ligand pocket that bridge 2 carbonyl groups of aspartame to residues D142 and L279. These results shed light on the activation mechanism and how signal transmission arising from the extracellular domain of the T1R2 monomer of the sweet receptor leads to the perception of sweet taste.

The anatomy of mammalian sweet taste receptors

“…In the model of T1R2, the residue S40 is located in this cavity. This finding is in line with the mutation S40A that decreases the response to aspartame . As with mGluR, this second pocket offers an opportunity to design new potent sweeteners.…”

“…A refined alignment was obtained by checking that the available site‐directed mutagenesis in vitro data for all these receptors were fitting the model (Table ). One can note a minor difference with already published sequence alignments for the residue K65 in T1R2 consistent with a putative role in the maintain of a closed structure of the VFD . The alignment unveils a moderate degree of sequence identity within the class C sub‐family (29 ± 1% identity between T1Rs and mGluRs and 39% identity between T1R2 and T1R3, cf.…”

“…S2 and S4) or based on the homology model are written in italic. Amino‐acids involved in the VFD orthosteric binding site (in blue) correspond to the Wellendorph and Bräuner‐Osborne's definition of the orthosteric binding site homology of class C GPCRs and mutagenesis studies of Xu et al ., Proc Natl Acad Sci, 2004; Zhang et al ., Proc Natl Acad Sci, 2010; Masuda et al ., Plos One, 2012; Maillet et al ., Chem Senses, 2015 . Residues of the allosteric binding sites (in green) come from experimental studies of T1R3 by Jiang et al ., J Biol Chem, 2005 and Winnig et al ., BMC Struct Biol, 2007 and then extrapolated to T1R2.…”

“…Typically, in the field of chemical senses, ligand–receptor interactions and dynamics were discussed on G protein‐coupled odorant receptors . More specifically, considering taste receptors, models of the sweet taste VFD and TMD have been proposed based on other class C VFD and on class A GPCR templates, respectively.…”

The cephalic phase insulin response to nutritive and low-calorie sweeteners in solid and beverage form

“…Whether this binding site holds special importance for the CPIR awaits further study. Aspartame is one low calorie sweetener that has not been associated with a CPIR or biphasic response in humans [51] and it binds to the amino acid terminal domain (ATD) of the T1R2 component of the sweet receptor [54, 57]. Activation of sweet receptors on pancreatic beta cells by different sweeteners elicits considerable intracellular signaling specificity [58–60] consistent with potential differential downstream effects between low-calorie sweeteners.…”