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This article presents the results of a comprehensive toxicity assessment of brazzein and monellin, yeast-produced recombinant sweet-tasting proteins. Excessive sugar consumption is one of the leading dietary and nutritional problems in the world, resulting in health complications such as obesity, high blood pressure, and cardiovascular disease. Although artificial small-molecule sweeteners widely replace sugar in food, their safety and long-term health effects remain debatable. Many sweet-tasting proteins, including thaumatin, miraculin, pentadin, curculin, mabinlin, brazzein, and monellin have been found in tropical plants. These proteins, such as brazzein and monellin, are thousands-fold sweeter than sucrose. Multiple reports have presented preparations of recombinant sweet-tasting proteins. A thorough and comprehensive assessment of their toxicity and safety is necessary to introduce and apply sweet-tasting proteins in the food industry. We experimentally assessed acute, subchronic, and chronic toxicity effects, as well as allergenic and mutagenic properties of recombinant brazzein and monellin. Our study was performed on three mammalian species (mice, rats, and guinea pigs). Assessment of animals' physiological, biochemical, hematological, morphological, and behavioral indices allows us to assert that monellin and brazzein are safe and nontoxic for the mammalian organism, which opens vast opportunities for their application in the food industry as sugar alternatives. [ABSTRACT FROM AUTHOR]

This article presents the results of a comprehensive toxicity assessment of brazzein and monellin, yeast-produced recombinant sweet-tasting proteins. Excessive sugar consumption is one of the leading dietary and nutritional problems in the world, resulting in health complications such as obesity, high blood pressure, and cardiovascular disease. Although artificial small-molecule sweeteners widely replace sugar in food, their safety and long-term health effects remain debatable. Many sweet-tasting proteins, including thaumatin, miraculin, pentadin, curculin, mabinlin, brazzein, and monellin have been found in tropical plants. These proteins, such as brazzein and monellin, are thousands-fold sweeter than sucrose. Multiple reports have presented preparations of recombinant sweet-tasting proteins. A thorough and comprehensive assessment of their toxicity and safety is necessary to introduce and apply sweet-tasting proteins in the food industry. We experimentally assessed acute, subchronic, and chronic toxicity effects, as well as allergenic and mutagenic properties of recombinant brazzein and monellin. Our study was performed on three mammalian species (mice, rats, and guinea pigs). Assessment of animals’ physiological, biochemical, hematological, morphological, and behavioral indices allows us to assert that monellin and brazzein are safe and nontoxic for the mammalian organism, which opens vast opportunities for their application in the food industry as sugar alternatives.

• Enhancing the thermal stability of single-chain monellin (MNEI) via PyRosetta. • Four variants with T m > 96 °C while maintaining sweetness. • Variants remained soluble after boiling for one hour. • Variants displayed stability across alkaline, acidic, and neutral environments. • Potential food industry applications and valuable protein engineering strategies. Sweet proteins offer a promising solution as sugar substitutes by providing a sugar-like sweetness without the negative health impacts linked to sugar or artificial sweeteners. However, the low thermal stability of sweet proteins has hindered their applications. In this study, we took a computational approach utilizing ΔΔG calculations in PyRosetta to enhance the thermostability of single-chain monellin (MNEI). By generating and characterizing 21 variants with single mutation, we identified 11 variants with higher melting temperature (T m) than that of MNEI. To further enhance the thermal stability, we conducted structural analysis and designed an additional set of 14 variants with multiple mutations. Among these variants, four exhibited a significant improvement in thermal stability, with an increase of at least 20 °C (T m > 96 °C) compared to MNEI, while maintaining their sweetness. Remarkably, these variants remained soluble even after being heated in boiling water for one hour. Moreover, they displayed exceptional stability across alkaline, acidic and neutral environments. These findings highlight the tremendous potential of these variants for applications in the food and beverage industry. Additionally, this study provides valuable strategies for protein engineering to enhance the thermal stability of sweet proteins. [ABSTRACT FROM AUTHOR]

The plants described in this review are a source of natural high-intensity sweeteners, which can be used in food and by the pharmaceutical industry in the future. Most of the plants are still not approved for use, even though they are traditionally used in countries where they appear naturally. Ten of the herein described intense sweeteners are characterized by a much higher sweetness in relation to sucrose. The highest values were received for miraculin, obtained from Synsepalum dulcificum (400,000 times sweeter than sucrose, induced by citric acid); thaumatin (1,600 to 3,000 times sweeter), monatin (1,200 - 3,000) and pentadin (500 to 2,000 times sweeter). Some of these substances can also modify the taste, like changing sour into sweet taste (miraculin and neoculin). The most widely used sweeteners are steviol glycosides and thaumatin, which have been admitted for use as a sweetener in the European Union, while in the US, they have the GRAS status (thaumatin as a food enhancer). Mogroside obtained from Siraitia grosvenorii (called Luo Han Guo) is not approved for use in the EU, but was granted GRAS status in the US by the FDA. This gives a chance that it will soon be approved as a novel food or food additive in the European Union. [ABSTRACT FROM AUTHOR]

A few proteins, discovered mainly in tropical fruits, have a distinct sweet taste. These proteins have played an important role towards a molecular understanding of the mechanisms of taste. Owing to the huge difference in size, between most sweeteners and sweet proteins, it was believed that they must interact with a different receptor from that of small molecular weight sweeteners. Recent modelling studies have shown that the single sweet taste receptor has multiple active sites and that the mechanism of interaction of sweet proteins is intrinsically different from that of small sweeteners. Small molecular weight sweeteners occupy small receptor cavities inside two subdomains of the receptor, whereas sweet proteins can interact with the sweet receptor according to a mechanism called the ‘wedge model’ in which they bind to a large external cavity. This review describes these mechanisms and outlines a history of sweet proteins. [ABSTRACT FROM AUTHOR]