Large-scale production of recombinant miraculin protein in transgenic carrot callus supension cultures using air-lift bioreactors

Abstract

Miraculin, derived from the miracle fruit ( Synsepalum dulcificum ), is a taste-regulating protein that interacts with human sweet-taste receptors and transforms sourness into sweet taste. Since miracle fruit is cultivated in West Africa, mass production of miraculin is limited by regional and seasonal constraints. Here, we investigated mass production of recombinant miraculin in carrot ( Daucus carota L.) callus cultures using an air-lift bioreactor. To increase miraculin expression, the oxidative stress-inducible SWAP2 promoter was used to drive the expression of miraculin gene under various stress treatments. An 8 h treatment of hydrogen peroxide (H 2 O 2 ) and salt (NaCl) increased the expression of miraculin gene by 5-fold compared with the untreated control. On the other hand, abscisic acid, salicylic acid, and methyl jasmonate treatments showed no significant impact on miraculin gene expression compared with the control. This shows that since H 2 O 2 and NaCl treatments induce oxidative stress, they activate the SWPA2 promoter and consequently up-regulate miraculin gene expression. Thus, the results of this study provide a foundation for industrial-scale production of recombinant miraculin protein using transgenic carrot cells as a heterologous host.

Production of miraculin protein in suspension cell lines of transgenic rice usingAgrobacterium

“…최근 들어 miraculin 기능을 산업용 또는 재조합 miraculin 생산을 위해 대장균 (Kurihara 1992), 효모 (Kurihara 1997) 및 Aspergillus oryzae (Ito et al 2007)와 같은 다양한 숙주세포를 이용하여 실험한 결과 미라클 프루트 열매로부터 얻은 활성보다 훨씬 낮은 결과를 초래하였다. 반면에 식물 발현시스템을 통한 몇몇 실험에서 는 재조합 miraculin 활성과 천연 miraculin활성과 거의 비슷 한 결과를 얻었다 (Sun et al 2006, Sun et al 2007, Al Bachchu et al 2011, Park et al 2020. 이런 결과는 N-glycosylation 뿐만 아 니라 miraculin에서 β1,2-xylose (Xyl)과 α1,3-fucose (Fuc)잔기 들을 가진 식물 특이적 N-glycans이 단백질 접힘(folding), 안 정성 및 그에 따른 활성 등에 매우 중요하다고 하였다 (Kajiura et al 2018…”