이상엽 (KAIST)

한빛사논문

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KAIST

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Nat. Commun., Aug 24 2023, 14 (1) 5177 | https://doi.org/10.1038/s41467-023-40932-4 A synthetic cell-free 36-enzyme reaction system for vitamin B12 production.

Qian Kang^1,2,3, Huan Fang^1,2,3, Mengjie Xiang^1,2, Kaixing Xiao^2,3, Pingtao Jiang^2,3, Chun You^1,2, Sang Yup Lee⁴ & Dawei Zhang^1,2,3

¹University of Chinese Academy of Sciences, No.19 (A) Yuquan Road, Shijingshan District, 100049 Beijing, China.

²Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, 32 Xi Qi Dao, Tianjin Airport Economic Area, 300308 Tianjin, China.

³Key Laboratory of Engineering Biology for Low-Carbon Manufacturing, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, 300308 Tianjin, China.

⁴Department of Chemical and Biomolecular Engineering (BK21 four program), Korea Advanced Institute of Science and Technology (KAIST), 291 Daehak-ro, Yuseong-gu, Daejeon 34141, Republic of Korea.

Corresponding authors : Correspondence to Sang Yup Lee or Dawei Zhang.

Abstract

Adenosylcobalamin (AdoCbl), a biologically active form of vitamin B ₁₂ (coenzyme B ₁₂), is one of the most complex metal-containing natural compounds and an essential vitamin for animals. However, AdoCbl can only be de novo synthesized by prokaryotes, and its industrial manufacturing to date was limited to bacterial fermentation. Here, we report a method for the synthesis of AdoCbl based on a cell-free reaction system performing a cascade of catalytic reactions from 5-aminolevulinic acid (5-ALA), an inexpensive compound. More than 30 biocatalytic reactions are integrated and optimized to achieve the complete cell-free synthesis of AdoCbl, after overcoming feedback inhibition, the complicated detection, instability of intermediate products, as well as imbalance and competition of cofactors. In the end, this cell-free system produces 417.41 μg/L and 5.78 mg/L of AdoCbl using 5-ALA and the purified intermediate product hydrogenobyrate as substrates, respectively. The strategies of coordinating synthetic modules of complex cell-free system describe here will be generally useful for developing cell-free platforms to produce complex natural compounds with long and complicated biosynthetic pathways.

논문정보

형식Research article
게재일2023년 08월 (BRIC 등록일 2023-09-06)
연구진국내(교신)+국외 연구진
분야 생명과학 > 응용생물화학

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