Abstract: In this study,the codons of Mabinlin Ⅱ(chainA+chainB)were optimized according to the codon usage bias in E. coli.The new sweet protein gene was named MabinlinII(AB)Plus. The MabinlinⅡ(AB)Plus gene was cloned into the E. coli expression vector pET-28a(+). The recombinant vector pET-28a(+)-MabinlinⅡ(AB)Plus was then transferred into E. coli BL21(DE3)host cells. Induction by IPTG,the optimized gene MabinlinⅡ(AB)Plus could be highly expressed in E. coli. The results showed that the optimal expression condition of the codon-optimized gene MabinlinⅡ(AB)Plus in E. coli was:IPTG concentration 1.4 mmol/L,inducer temperature was 37℃,optimum strain initial growth amount OD₆₀₀,induction time was 0.7 h.Under the optimal induction expression conditions,the expression of the target protein expressed for about 33.0% of the total protein,which was about 7% higher than that before optimization. The supernatant of the target protein was affinity purified and had a single band at 19 kDa. After sensory evaluation,the sweetness of the target protein was about 400 times that of the 2% standard sucrose solution. This study would provide a scientific theoretical basis for the development of a new sweetener based on the sweet protein MabinlinⅡ.