title: Complete purification and characterization of the taste-modifying protein, miraculin, from miracle fruit.
abstract: The taste-modifying protein, miraculin, has the unusual property of modifying a sour taste into a sweet taste. Previous attempts to isolate miraculin from deeply colored alkaline extracts of the miracle fruit were unsuccessful. We found that miraculin is extracted with 0.5 M NaCl solution. The extracted solution is colorless and shows the strong sweet-inducing activity. Miraculin was purified from the extracted solution by ammonium sulfate fractionation, CM-Sepharose ion-exchange chromatography, and concanavalin A-Sepharose affinity chromatography. The purified miraculin thus obtained gave a single sharp peak in reverse phase high performance liquid chromatography, indicating that it is highly pure. The sample also gave a single band having molecular weight 28,000 in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This value was much lower than the values reported previously (40,000-48,000). The amino acid composition of the purified miraculin was determined. Sequence analysis of the purified miraculin indicated that it is composed of a pure single polypeptide and identified 20 amino-terminal amino acids. The purified miraculin contained as much as 13.9% of sugars, which consisted of glucosamine, mannose, galactose, xylose, and fucose in a molar ratio of 3.03:3.00:0.69:0.96:2.12.
authors.affiliation: Department of Chemistry, Faculty of Education, Yokohama National University, Japan.
authors.forename: S
authors.initials: S
authors.lastname: Theerasilp
authors.forename: Y
authors.initials: Y
authors.lastname: Kurihara
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D005638:Fruit / Q000032:analysis*; D006023:Glycoproteins / Q000302:isolation & purification* / Q000378:metabolism; D008970:Molecular Weight; D013549:Sweetening Agents / Q000302:isolation & purification* / Q000378:metabolism
chemical_list: D000596:Amino Acids; D006023:Glycoproteins; D013549:Sweetening Agents; C008533:miraculin protein, Synsepalum dulcificum