Improved expression of recombinant sweet-tasting brazzein using codon optimization and host change as new strategies

• Published in: Food biotechnology Vol. 34; no. 1; pp. 62 - 76
• Main Authors: Jafarian, Vahab, Bagheri, Khadijeh, Zarei, Jabraeil, Karami, Shima, Ghanavatian, Parisa
• Format: Journal Article
• Language: English
• Published: Philadelphia Taylor & Francis 02-01-2020; Taylor & Francis Ltd
• Subjects: BL21(DE3); Disulfide bonds; E coli; Food industry; Food processing industry; GC content; High temperature; natural sweetener; Optimization; Proteins; recombinant protein; SHuffle® T7 Express; Sucrose; Sugar; Sweet taste; Sweeteners; Sweetness
• DOI: 10.1080/08905436.2019.1711113
• EISSN: 1532-4249
• ISSN: 0890-5436

There is increased food industry attention to low-calorie natural sweeteners with good taste properties such as in plant-based sweet proteins. Brazzein is an attractive alternative to sucrose because of its intense sweetness, natural origin, and good stability at wide pH ranges and high temperature. In the present study, by using the minor form of brazzein as the basic sequence, the codon-optimized brazzein gene was designed based on appropriate GC content and preferred codon of E. coli. Then, the synthesized brazzein gene was cloned in pET28a vector and expressed in BL21(DE3) and SHuffle® T7 Express strains. Recombinant brazzein was expressed about 1.8 to 2.3 mg/L and 7.2 to 8.4 mg/L in BL21 (DE3) and SHuffle® T7 Express strain, respectively. Recombinant brazzein with His-tag lacked the sweetness. After the removal of the His-tag, it was shown that in addition to sweetness potency, the amount of brazzein proteins purified from SHuffle® T7 Express was higher than that from BL21. This improvement could be due to the better performance of SHuffle® T7 Express in the expression of the protein with high disulfide bridges and the effect of disulfide bonds on the sweetness of brazzein.