The application discloses a brazzein
mutant and application thereof, and belongs to the technical field of
protein modification. In view of the defects that wild-type brazzein is insufficient in
sweetness, difficult to meet the application demand of industrialized
sugar substitute, and the yield of a microbial
heterologous expression
system needs to be improved, the application adopts
protein engineering directed evolution technology, screens single mutants P13E, D26K and I49G, and further constructs combination mutants P13E / D26K, P13E / I49G, D26K / I49G and P13E / D26K / I49G. The mutants are heterogeneously and efficiently expressed in a
Pichia pastoris expression
system, and the
sweet taste performance of the mutants is systematically evaluated. The results show that, compared with the wild-type brazzein, the
sweetness of each
mutant is increased by 60% to 340%, and the mutants have excellent stability and pure
sweet taste, and can be used as a new type of sweetener and applied to the field of
food processing, and have important industrial application value.