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The sweet protein monellin has high sweet potency with limited stability. In this study, 3 double-sites mutants (E2N/E23A, E2N/Y65R and E23A/Y65R) of the single-chain monellin (MNEI) were constructed. The proteins were expressed in E. coli BL21 and purified to homogeneity by nickel affinity chromatography with yields above 10 mg/L cell culture. Introduction of a sweeter mutant E2N into E23A or Y65R...
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