The primary structure of the sweet-tasting protein thaumatin has been elucidated. The protein consists of a single polypeptide chain of 207 residues. The sequence of the N-terminal part of the chain was determined by sequenator analysis. As the protein contains only one methionine residue, it was possible to deduce the N-terminal sequence of the C-terminal cyanogen bromide fragment by automatic sequencing of the cyanogen-bromide-cleaved, succinylated protein. To arrive at the sequence of the whole protein tryptic and Staphylococcus protease peptides, together with chymotryptic peptides and a 2-(2-nitrophenylsulfenyl)-3-methyl-3'-bromoindolenine (BNPS-skatole) fragment were also sequenced.Comparing the amino acid sequence of thaumatin with that of the other sweet-tasting protein, monellin, we have located five sets of identical tripeptides. Since immunological cross-reactivity of thaumatin antibodies with monellin has recently been described, one or more of these tripeptides might be part of a common antibody recombination site and possibly be involved in the interaction with the sweet-taste receptor.

From an aqueous extract of the fruit of the tropical plant Thaumatococcus ohniellii &frtfh, -two sweet-tasting basic proteins, here named thaumatin I and thaumatin 11, were isolated by ultrafiltration, gel filtration on Sephadex (2-50 and ion-exchange chromatography on SE-Sephadex C-25, using a sodium chloride concentration gradient.The proteinaceous character of the two sweet-tasting compounds was proven by the characteristic ultraviolet absorption spectrum, the presence of almost 100 polypeptide material as determined by the biuret method, the yield of 100°/o amino acids on hydrolysis with hydrochlorid acid, the positive reaction with amido black colouring agent (as used in the analysis of the fractions by polyacrylamide gel and starch gel electrophoresis) and by the disappearance of the sweet taste of the compounds after incubation with trypsin.The basic character of the two proteins appears from their isoelectric points of almost 12, as estimated by starch gel electrophoresis a t M e r e n t p H values, and from the precipitation of the proteins a t pH 12.

Thaumatin and monellin are the two sweetest compounds known to man-about 100,000 times sweeter than sugar on a molar basis and 3000 times on a weight basis. These proteins represent a unique class of proteins that are tasteactive. We report the three-dimensional structure of thaumatin I at 3.1 A resolution.Taste is one of the least understood senses from a biochemical point of view. Sweet taste can be elicited to varying degrees by a wide variety of compounds in high concentration, such as mono-and disaccharides, several amino acids, dipeptide derivatives, glycerol, cyclamates, saccharine, acesulfame, some hydrated inorganic compounds, and others.Sensations of sweet taste are elicited by sweet compounds binding to sweet receptors of taste buds on the tongue, similar to hormone-hormone receptor binding. However, the similarity ends here. A 50% binding of hormone receptors usually occurs at around 10-8 to 10-11 M concentration of hormones, but in the case of sweet receptors the concentration to register sweet taste is about 10-1 to 10-3 M of sucrose. Furthermore, like other sensory perceptions, taste is associated with a unique set of sensation parameters. These are threshold concentration, onset time, intensity, saturation concentration, lingering time, aftertaste, and potentiation of other tastes or flavors. Systematic studies of these properties of taste-active compounds have been difficult in the past because of high levels of nonspecific binding between the sweet comnpounds and the sweet receptors due to the high concentrations of the sweet compounds required to elicit the sweet taste.One of the most interesting and unusual recent findings is the existence of two intensely sweet proteins, monellin and thaumatin (for a recent review, see ref. 1). The sweet taste of the proteins can be registered at a very low concentration (10' M), comparable to those in hormone-hormone receptor interaction. These proteins are about 100,000 times sweeter than sucrose on a molar basis and several thousand times sweeter on a weight basis. In fact, these two proteins are the two sweetest compounds known to man. With these proteins, studies similar to those performed for hormonehormone receptors can be initiated. Three-dimensional structures of these proteins are likely to provide a solid foundation upon which studies to understand the molecular basis of sweet taste can be designed. We here report the crystal structure of thaumatin I at 3.1 A resolution.Thaumatins have been isolated from the fruit of a West African rain forest shrub, Thaumatococcus daniellii Benth (2), which has been used for centuries by inhabitants of the region to sweeten foods such as bread and palm wine, There are two major sweet proteins in the fruit, thaumatin I and II, with almost identical molecular weights of 22,000. They consist of 207 amino acids and have identical amino acid sequences except for five residues (3, 4). There are no histidine residues, but there are eight disulfide bonds. Partially pure thaumatin is available commercially from Si...

SUMMARY1. The gustatory effects of miraculin, the sweetness-inducing protein from the miracle fruit Synsepalum dulcificum, was studied in the rhesus monkey, Macaca mulatta.2. The intake of five acids was recorded in two-bottle preference tests, one bottle containing acid and the other tap water, before and after miraculin treatment. All the acids tasted more pleasant after miraculin.3. The electrical activity of the chorda tympani nerve to stimulation of the tongue with a variety of sweeteners, acids, sodium chloride and quinine hydrochloride was recorded in anaesthetized animals.4. Pre-treatment of the tongue with 03-5 mg miraculin doubled the summated nerve response to the acids and diminished the response to sucrose by about 10 %. The enhancement lasted for at least an hour and the diminution up to 20 min.5. After miraculin treatment the Spearman's rank correlation coefficient between the order of increased intake of acids and the order of enhancement of the summated nerve response was 0 99. 6. A solution of 0-1 mg miraculin per ml. elicited a weak nerve response. No preference over water for this concentration of miraculin was recorded in the twobottle tests.7. The activity of twenty-nine single taste fibres, selected for their responsiveness to sweetness or acids or both, was recorded after miraculin treatment. Effects were obtained in nine fibres which were similar but more pronounced than those observed in the summated recordings. Before miraculin, these fibres responded better and to a larger variety ofsweeteners (81 %) than the other fibres (40 %). After miraculin, acids elicited on the average 2-3 times more activity than before, while the response to sweeteners was depressed. In twenty fibres no effect of miraculin was observed. These fibres responded to fewer ofthe sweeteners and were more stimulated by the non-sweet stimuli than the first group.The authors' names are in alphabetical order.