Expression of a triple mutational des‐pGlu brazzein in transgenic mouse milk

Brazzein has excellent potential for use as a sweetener because of its high level of sweetening potency and stability against extreme temperature and pH. It is extracted from the tropical and difficult—to‐cultivate African plant Pentadiplandra brazzeana, which hampers its commercial viability. Here we report the mammary‐specific expression of wildtype or triple mutational (H31R/E36D/E41A) des‐pGlu brazzeins in the milk of transgenic mice. Using enzyme‐linked immunoassay (ELISA), western blot, and sweetness intensity testing, we confirmed that the triple mutation made the des‐pGlu brazzein molecule 10,000 times sweeter than sucrose in a weight base, even after 10 min of incubation at 100 °C; in addition, the triple mutant was also significantly sweeter than the wildtype des‐pGlu brazzein. This study provides new insights for producing brazzein or brazzein‐sweetened milk from animals for use in food and healthcare applications.