Elucidation of the structure-function relationships in sweet-tasting proteins by x-ray analysis at atomic resolution

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The crystal structure of a recombinant thaumatin I was determined to a resolution of 1.1 A (PDB:3AL7). The crystallographic structure of one of its variants, thaumatin II, was determined at a resolution of 1.27 A (PDB:3AOK). The results showed the critical positive charges are disordered and the flexibility and fluctuation of these side chains would be suitable for interaction with sweet receptors. The atomic structure of a recombinant thaumatin at pH 8.0 was determined at a resolution of 1.0 A (PDB:3VJQ). The results suggested a striking increase in the mobility of some lysine residues, which could facilitate a reaction with a free sulfhydryl residue produced via the β-elimination of disulfide bonds by heating at a pH above 7.0. Various mutants of thaumatin were prepared and their sweetness were quantitatively evaluated by cell-based assays using HEK293 cells expressing human sweet receptors. Chimeric human- mouse sweet receptors were constructed and their responses to sweeteners were investigated. The results showed the cysteine-rich domain of human T1R3 is important for the response to thaumatin.