Direct NMR detection of the binding of functional ligands to the human sweet receptor, a heterodimeric family 3 GPCR

Fariba M. Assadi-Porter; Marco Tonelli; Emeline Maillet; Klaas Hallenga; Outhiriaradjou Benard; Marianna Max; John L. Markley

doi:10.1021/ja8016939

Direct NMR detection of the binding of functional ligands to the human sweet receptor, a heterodimeric family 3 GPCR

Fariba M. Assadi-Porter, Marco Tonelli, Emeline Maillet, Klaas Hallenga, Outhiriaradjou Benard, Marianna Max, John L. Markley

Research output: Contribution to journal › Article › peer-review

63 Scopus citations

Abstract

We present a robust method for monitoring the binding of ligands to the heterodimeric (T1R2+T1R3) human sweet receptor (a family 3 GPCR receptor). The approach utilizes saturation transfer difference (STD) NMR spectroscopy with receptor proteins expressed on the surface of human epithelial kidney cells. The preparation investigated by NMR can contain either live cells or membranes isolated from these cells containing the receptor. We have used this approach to confirm the noncompetitive binding of alitame and cyclamate to the receptor and to determine that greatly reduced receptor binding affinity compared to wild-type brazzein explains the lack of sweetness of brazzein mutant A16C17. This approach opens new avenues for research on the mechanism of action of the sweet receptor and for the design of new noncalorigenic sweeteners.

Original language	English (US)
Pages (from-to)	7212-7213
Number of pages	2
Journal	Journal of the American Chemical Society
Volume	130
Issue number	23
DOIs	https://doi.org/10.1021/ja8016939
State	Published - Jun 11 2008
Externally published	Yes

ASJC Scopus subject areas

Catalysis
General Chemistry
Biochemistry
Colloid and Surface Chemistry

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abstract = "We present a robust method for monitoring the binding of ligands to the heterodimeric (T1R2+T1R3) human sweet receptor (a family 3 GPCR receptor). The approach utilizes saturation transfer difference (STD) NMR spectroscopy with receptor proteins expressed on the surface of human epithelial kidney cells. The preparation investigated by NMR can contain either live cells or membranes isolated from these cells containing the receptor. We have used this approach to confirm the noncompetitive binding of alitame and cyclamate to the receptor and to determine that greatly reduced receptor binding affinity compared to wild-type brazzein explains the lack of sweetness of brazzein mutant A16C17. This approach opens new avenues for research on the mechanism of action of the sweet receptor and for the design of new noncalorigenic sweeteners.",

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AU - Assadi-Porter, Fariba M.

AU - Tonelli, Marco

AU - Maillet, Emeline

AU - Hallenga, Klaas

AU - Benard, Outhiriaradjou

AU - Max, Marianna

AU - Markley, John L.

PY - 2008/6/11

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Direct NMR detection of the binding of functional ligands to the human sweet receptor, a heterodimeric family 3 GPCR

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