title: A Hypersweet Protein: Removal of The Specific Negative Charge at Asp21 Enhances Thaumatin Sweetness.
abstract: Thaumatin is an intensely sweet-tasting protein that elicits sweet taste at a concentration of 50 nM, a value 100,000 times larger than that of sucrose on a molar basis. Here we attempted to produce a protein with enhanced sweetness by removing negative charges on the interacting side of thaumatin with the taste receptor. We obtained a D21N mutant which, with a threshold value 31 nM is much sweeter than wild type thaumatin and, together with the Y65R mutant of single chain monellin, one of the two sweetest proteins known so far. The complex model between the T1R2-T1R3 sweet receptor and thaumatin, derived from tethered docking in the framework of the wedge model, confirmed that each of the positively charged residues critical for sweetness is close to a receptor residue of opposite charge to yield optimal electrostatic interaction. Furthermore, the distance between D21 and its possible counterpart D433 (located on the T1R2 protomer of the receptor) is safely large to avoid electrostatic repulsion but, at the same time, amenable to a closer approach if D21 is mutated into the corresponding asparagine. These findings clearly confirm the importance of electrostatic potentials in the interaction of thaumatin with the sweet receptor.
authors.affiliation: Laboratory of Food and Environmental Science, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011, Japan.
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Masuda
authors.affiliation: Laboratory of Food and Environmental Science, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011, Japan.
authors.forename: Keisuke
authors.initials: K
authors.lastname: Ohta
authors.affiliation: Laboratory of Food and Environmental Science, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011, Japan.
authors.forename: Naoko
authors.initials: N
authors.lastname: Ojiro
authors.affiliation: Laboratory of Food and Environmental Science, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011, Japan.
authors.forename: Kazuki
authors.initials: K
authors.lastname: Murata
authors.affiliation: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011, Japan.
authors.forename: Bunzo
authors.initials: B
authors.lastname: Mikami
authors.affiliation: Laboratory of Food and Environmental Science, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011, Japan.
authors.forename: Fumito
authors.initials: F
authors.lastname: Tani
authors.affiliation: Department of Basic and Clinical Neurosciences, King's College, London, London SE59RX (UK).|Dipartimento di Chimica, Universita' di Napoli Federico II, Napoli, I-80126, Italy.
authors.forename: Piero Andrea
authors.initials: PA
authors.lastname: Temussi
authors.affiliation: Department of Foods and Human Nutrition, Notre Dame Seishin University, Okayama 700-8516, Japan.
authors.forename: Naofumi
authors.initials: N
authors.lastname: Kitabatake
mesh_terms: D001216:Asparagine / Q000235:genetics*; D001665:Binding Sites; D008958:Models, Molecular; D062105:Molecular Docking Simulation; D009154:Mutation; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D055672:Static Electricity; D013649:Taste; D019862:Zingiberales / Q000235:genetics / Q000378:metabolism*
chemical_list: D010940:Plant Proteins; D001216:Asparagine
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