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The binding site for neohesperidin dihydrochalcone at the human sweet taste receptor.

Winnig M, Bufe B, Kratochwil NA, Slack JP, Meyerhof W - BMC Struct. Biol. (2007)

Bottom Line: From our data we conclude that we identified the neohesperidin dihydrochalcone binding site at the human sweet taste receptor, which overlaps with those for the sweetener cyclamate and the sweet taste inhibitor lactisole.This readily delivers a molecular explanation of our finding that lactisole is a competitive inhibitor of the receptor activation by neohesperidin dihydrochalcone and cyclamate.Some of the amino acid positions crucial for activation of hTAS1R2+hTAS1R3 by neohesperidin dihydrochalcone are involved in the binding of allosteric modulators in other class C GPCRs, suggesting a general role of these amino acid positions in allosterism and pointing to a common architecture of the heptahelical domains of class C GPCRs.

View Article: PubMed Central - HTML - PubMed

Affiliation: German Institute of Human Nutrition Potsdam-Rehbruecke, Department of Molecular Genetics, Arthur-Scheunert Allee 114-116, 14558 Nuthetal, Germany. marcel.winnig.mw@axxam.com

ABSTRACT

Background: Differences in sweet taste perception among species depend on structural variations of the sweet taste receptor. The commercially used isovanillyl sweetener neohesperidin dihydrochalcone activates the human but not the rat sweet receptor TAS1R2+TAS1R3. Analysis of interspecies combinations and chimeras of rat and human TAS1R2+TAS1R3 suggested that the heptahelical domain of human TAS1R3 is crucial for the activation of the sweet receptor by neohesperidin dihydrochalcone.

Results: By mutational analysis combined with functional studies and molecular modeling we identified a set of different amino acid residues within the heptahelical domain of human TAS1R3 that forms the neohesperidin dihydrochalcone binding pocket. Sixteen amino acid residues in the transmembrane domains 2 to 7 and one in the extracellular loop 2 of hTAS1R3 influenced the receptor's response to neohesperidin dihydrochalcone. Some of these seventeen residues are also part of the binding sites for the sweetener cyclamate or the sweet taste inhibitor lactisole. In line with this observation, lactisole inhibited activation of the sweet receptor by neohesperidin dihydrochalcone and cyclamate competitively, whereas receptor activation by aspartame, a sweetener known to bind to the N-terminal domain of TAS1R2, was allosterically inhibited. Seven of the amino acid positions crucial for activation of hTAS1R2+hTAS1R3 by neohesperidin dihydrochalcone are thought to play a role in the binding of allosteric modulators of other class C GPCRs, further supporting our model of the neohesperidin dihydrochalcone pharmacophore.

Conclusion: From our data we conclude that we identified the neohesperidin dihydrochalcone binding site at the human sweet taste receptor, which overlaps with those for the sweetener cyclamate and the sweet taste inhibitor lactisole. This readily delivers a molecular explanation of our finding that lactisole is a competitive inhibitor of the receptor activation by neohesperidin dihydrochalcone and cyclamate. Some of the amino acid positions crucial for activation of hTAS1R2+hTAS1R3 by neohesperidin dihydrochalcone are involved in the binding of allosteric modulators in other class C GPCRs, suggesting a general role of these amino acid positions in allosterism and pointing to a common architecture of the heptahelical domains of class C GPCRs.

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Inhibition of hTAS1R2/hTAS1R3 by lactisole. (A-D) Concentration responses of cells transfected with hTAS1R2-hTAS1R3 DNAs to sweetners mixed with different lactisole concentrations. No lactisole present (filled circles, solid line), 50 μM lactisole (filled triangles up, dashed line), and 100 μM lactisole (filled squares, dash-dotted line). (E) EC50 values for acesulfame K, aspartame, NHDC, and cyclamate in the absence or presence of 50 μM and 100 μM lactisole.
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Figure 2: Inhibition of hTAS1R2/hTAS1R3 by lactisole. (A-D) Concentration responses of cells transfected with hTAS1R2-hTAS1R3 DNAs to sweetners mixed with different lactisole concentrations. No lactisole present (filled circles, solid line), 50 μM lactisole (filled triangles up, dashed line), and 100 μM lactisole (filled squares, dash-dotted line). (E) EC50 values for acesulfame K, aspartame, NHDC, and cyclamate in the absence or presence of 50 μM and 100 μM lactisole.

Mentions: Recently, it has been shown that the binding sites for the sweetener cyclamate and the sweet inhibitor lactisole overlap in the heptahelical domain of TAS1R3 [14,15]. We therefore reasoned that the NHDC binding site might also overlap with that for lactisole. In this case one would expect that lactisole competitively inhibits the activation of hTAS1R2/hTAS1R3 by NHDC and cyclamate. In contrast, receptor activation by sweeteners that bind to other sites should be allosterically inhibited. To verify this assumption we recorded concentration-response curves for NHDC, cyclamate, acesulfame K and aspartame in the presence of different concentrations of lactisole in cells expressing hTAS1R2/hTAS1R3 (Fig. 2).


The binding site for neohesperidin dihydrochalcone at the human sweet taste receptor.

Winnig M, Bufe B, Kratochwil NA, Slack JP, Meyerhof W - BMC Struct. Biol. (2007)

Inhibition of hTAS1R2/hTAS1R3 by lactisole. (A-D) Concentration responses of cells transfected with hTAS1R2-hTAS1R3 DNAs to sweetners mixed with different lactisole concentrations. No lactisole present (filled circles, solid line), 50 μM lactisole (filled triangles up, dashed line), and 100 μM lactisole (filled squares, dash-dotted line). (E) EC50 values for acesulfame K, aspartame, NHDC, and cyclamate in the absence or presence of 50 μM and 100 μM lactisole.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2099433&req=5

Figure 2: Inhibition of hTAS1R2/hTAS1R3 by lactisole. (A-D) Concentration responses of cells transfected with hTAS1R2-hTAS1R3 DNAs to sweetners mixed with different lactisole concentrations. No lactisole present (filled circles, solid line), 50 μM lactisole (filled triangles up, dashed line), and 100 μM lactisole (filled squares, dash-dotted line). (E) EC50 values for acesulfame K, aspartame, NHDC, and cyclamate in the absence or presence of 50 μM and 100 μM lactisole.
Mentions: Recently, it has been shown that the binding sites for the sweetener cyclamate and the sweet inhibitor lactisole overlap in the heptahelical domain of TAS1R3 [14,15]. We therefore reasoned that the NHDC binding site might also overlap with that for lactisole. In this case one would expect that lactisole competitively inhibits the activation of hTAS1R2/hTAS1R3 by NHDC and cyclamate. In contrast, receptor activation by sweeteners that bind to other sites should be allosterically inhibited. To verify this assumption we recorded concentration-response curves for NHDC, cyclamate, acesulfame K and aspartame in the presence of different concentrations of lactisole in cells expressing hTAS1R2/hTAS1R3 (Fig. 2).

Bottom Line: From our data we conclude that we identified the neohesperidin dihydrochalcone binding site at the human sweet taste receptor, which overlaps with those for the sweetener cyclamate and the sweet taste inhibitor lactisole.This readily delivers a molecular explanation of our finding that lactisole is a competitive inhibitor of the receptor activation by neohesperidin dihydrochalcone and cyclamate.Some of the amino acid positions crucial for activation of hTAS1R2+hTAS1R3 by neohesperidin dihydrochalcone are involved in the binding of allosteric modulators in other class C GPCRs, suggesting a general role of these amino acid positions in allosterism and pointing to a common architecture of the heptahelical domains of class C GPCRs.

View Article: PubMed Central - HTML - PubMed

Affiliation: German Institute of Human Nutrition Potsdam-Rehbruecke, Department of Molecular Genetics, Arthur-Scheunert Allee 114-116, 14558 Nuthetal, Germany. marcel.winnig.mw@axxam.com

ABSTRACT

Background: Differences in sweet taste perception among species depend on structural variations of the sweet taste receptor. The commercially used isovanillyl sweetener neohesperidin dihydrochalcone activates the human but not the rat sweet receptor TAS1R2+TAS1R3. Analysis of interspecies combinations and chimeras of rat and human TAS1R2+TAS1R3 suggested that the heptahelical domain of human TAS1R3 is crucial for the activation of the sweet receptor by neohesperidin dihydrochalcone.

Results: By mutational analysis combined with functional studies and molecular modeling we identified a set of different amino acid residues within the heptahelical domain of human TAS1R3 that forms the neohesperidin dihydrochalcone binding pocket. Sixteen amino acid residues in the transmembrane domains 2 to 7 and one in the extracellular loop 2 of hTAS1R3 influenced the receptor's response to neohesperidin dihydrochalcone. Some of these seventeen residues are also part of the binding sites for the sweetener cyclamate or the sweet taste inhibitor lactisole. In line with this observation, lactisole inhibited activation of the sweet receptor by neohesperidin dihydrochalcone and cyclamate competitively, whereas receptor activation by aspartame, a sweetener known to bind to the N-terminal domain of TAS1R2, was allosterically inhibited. Seven of the amino acid positions crucial for activation of hTAS1R2+hTAS1R3 by neohesperidin dihydrochalcone are thought to play a role in the binding of allosteric modulators of other class C GPCRs, further supporting our model of the neohesperidin dihydrochalcone pharmacophore.

Conclusion: From our data we conclude that we identified the neohesperidin dihydrochalcone binding site at the human sweet taste receptor, which overlaps with those for the sweetener cyclamate and the sweet taste inhibitor lactisole. This readily delivers a molecular explanation of our finding that lactisole is a competitive inhibitor of the receptor activation by neohesperidin dihydrochalcone and cyclamate. Some of the amino acid positions crucial for activation of hTAS1R2+hTAS1R3 by neohesperidin dihydrochalcone are involved in the binding of allosteric modulators in other class C GPCRs, suggesting a general role of these amino acid positions in allosterism and pointing to a common architecture of the heptahelical domains of class C GPCRs.

Show MeSH
Related in: MedlinePlus