食品与发酵工业 >

2025 , Vol. 51 >Issue 21: 27 - 34

DOI: https://doi.org/10.13995/j.cnki.11-1802/ts.042062

研究报告

重组大肠杆菌全细胞转化反式茴脑合成茴香醛

  • 张志凯 ,
  • 林谦
展开
  • 1(玉林师范学院 生物与制药学院,广西 玉林,537000)
    2(玉林师范学院,广西高校山地生物多样性保育重点实验室,广西 玉林,537000)
第一作者：博士,副教授(林谦教授为通信作者,E-mail:linqian1977@163.com)

收稿日期: 2025-01-08

  修回日期: 2025-02-19

  网络出版日期: 2025-11-21

基金资助

广西自然科学基金项目(2022GXNSFAA035443);玉林师范学院博士启动基金项目(G2022ZK34)

收起

Biosynthesis of p-anisaldehyde from trans-anethole by whole-cell biotransformation using recombinant Escherichia coli

  • ZHANG Zhikai ,
  • LIN Qian
Expand
  • 1(College of Biology and Pharmacy, Yulin Normal University, Yulin 537000, China)
    2(Key Laboratory of Mountain Biodiversity Conservation, Education Department of Guangxi Zhuang Autonomous Region, Yulin Normal University, Yulin 537000, China)

Received date: 2025-01-08

  Revised date: 2025-02-19

  Online published: 2025-11-21

Fold

摘要

茴香醛是食品饮料、化妆品行业广泛应用的香料。为提高生物合成的产量和转化率,该文研究了茴香醛的全细胞催化合成。将Paraburkholderia sp.MR185的反式茴脑加氧酶与反向蛋白XXA在大肠杆菌中融合表达,将诱导后的菌体用作全细胞催化剂转化反式茴脑合成茴香醛,并制成固定化细胞。诱导后的重组菌检测到了反式茴脑加氧酶活性,裂解上清液及沉淀中均检测到了融合蛋白。诱导条件优化后,菌体酶活力达到9.3 U/g。全细胞催化条件优化后,转化液中茴香醛的产量达到1.62 g/L,摩尔转化率达到88.3%。用海藻酸钠包埋重组菌体细胞,将固定化细胞用于重复转化反式茴脑生成茴香醛的反应,得到的茴香醛产量达到1.26 g/L,摩尔转化率为68.5%,前3次的产量均达到1 g/L以上。该研究为首次应用大肠杆菌全细胞催化合成茴香醛,为茴香醛生物合成产业化提供了技术参考。

关键词： 茴香醛; 反式茴脑; 反式茴脑加氧酶; 大肠杆菌; 全细胞催化

本文引用格式

张志凯 , 林谦 . 重组大肠杆菌全细胞转化反式茴脑合成茴香醛[J]. 食品与发酵工业, 2025 , 51(21) : 27 -34 . DOI: 10.13995/j.cnki.11-1802/ts.042062

Abstract

p-Asnisaldehyde is an aromatic compound widely used in food, beverage, and cosmetics industries.To increase biosynthesis yield and conversion rate, whole-cell biocatalysis for p-anisaldehyde production was investigated.The trans-anethole oxygenase from Paraburkholderia sp.MR185 and a solubilization tag retro-protein XXA was fused and expressed in Escherichia coli BL21(DE3).The induced E.coli cells were used as whole-cell biocatalyst for conversion of trans-anethole to p-anisaldehyde, and were immobilized.The activity of trans-anethole oxygenase was dectected in induced E.coli cells and the fusion protein was present in the supernatant and sediment of cell extract.The induction conditions were optimized and the enzyme activity in E.coli cells reached 9.3 U/g.Under optimized conditions of whole-cell transformation, the yield of p-anisaldehyde reached 1.62 g/L with the conversion rate being 88.3%.E.coli cells were immobilized in sodium alginate and used for repeated transformation of trans-anethole to p-anisaldehyde.The yield of of p-anisaldehyde reached 1.26 g/L with the conversion rate being 68.5%, and in the fisrt 3 runs the yield of p-anisaldehyde were above 1 g/L.This study is the first report on whole-cell biocatalyst for p-anisaldehyde synthesis, and provides a technical reference for the industrialized biosynthesis of p-anisaldehyde.

Key words： p-anisaldehyde; trans-anethole; trans-anethole oxygenase; Escherichia coli; whole-cell biocatalysis

参考文献

[1] XIONG J, YUAN X, ZONG M H, et al.Iron-incorporated metal-organic frameworks for oxidative cleavage of trans-anethole to p-anisaldehyde[J].Nanoscale, 2023, 15(48):19493-19498.
[2] MANINA A S, FORLANI F.Biotechnologies in perfume manufacturing:Metabolic engineering of terpenoid biosynthesis[J].International Journal of Molecular Sciences, 2023, 24(9):7874.
[3] DICKEY R M, GOPAL M R, NAIN P, et al.Recent developments in enzymatic and microbial biosynthesis of flavor and fragrance molecules[J].Journal of Biotechnology, 2024, 389:43-60.
[4] HAN D F, RYU J Y, KANALY R A, et al.Isolation of a gene responsible for the oxidation of trans-anethole to para-anisaldehyde by Pseudomonas putida JYR-1 and its expression in Escherichia coli[J].Applied and Environmental Microbiology, 2012, 78(15):5238-5246.
[5] HAN D F, SADOWSKY M J, CHONG Y, et al.Characterization of a self-sufficient trans-anethole oxygenase from Pseudomonas putida JYR-1[J].PLoS One, 2013, 8(9):e73350.
[6] 粟桂娇, 刘雄民, 朱萍, 等.反式茴脑生物降解菌株的筛选及其生物转化[J].化工进展, 2009, 28(1):141-144;172.
SU G J, LIU X M, ZHU P, et al.Isolation of trans-anethole degrading bacterial strains and biotransformation of trans-anethole[J].Chemical Industry and Engineering Progress, 2009, 28(1):141-144;172.
[7] 粟桂娇, 刘雄民, 李伟光, 等.两相系统中生物转化茴脑生成茴香醛[J].精细化工, 2009, 26(2):136-141.
SU G J, LIU X M, LI W G, et al.Biotransformation of anethole to anisaldehyde in aqueous-organic solvents biphasic systems[J].Fine Chemicals, 2009, 26(2):136-141.
[8] 梁肖仍, 宋张杨, 曾华贺, 等.一株转化茴脑生成茴香醛的菌株筛选与鉴定[J].食品与生物技术学报, 2011, 30(6):956-961.
LIANG X R, SONG Z Y, ZENG H H, et al.Isolation and characterization of a strain for bioconversion of anethole to anialdehyde[J].Journal of Food Science and Biotechnology, 2011, 30(6):956-961.
[9] 刘思琴, 郑璞, 陈鹏程, 等.合成洋茉莉醛重组菌株的构建与产物鉴定[J].食品与发酵工业, 2018, 44(6):1-7.
LIU S Q, ZHENG P, CHEN P C, et al.Construction of a recombinant strain for biosynthesis of heliotropin and its product identification[J].Food and Fermentation Industries, 2018, 44(6):1-7.
[10] WEN P, WU D, ZHENG P, et al.Highly efficient biosynthesis of heliotropin by engineered Escherichia coli coexpressing trans-anethole oxygenase and formate dehydrogenase[J].Journal of Agricultural and Food Chemistry, 2019, 67(51):14121-14128.
[11] LIN Q, LI J N, LING X R, et al.Cloning and expression of a novel trans-anethole oxygenase gene from Paraburkholderia sp.MR185[J].Journal of General and Applied Microbiology, 2022, 68(3):163-167.
[12] PURWANI N N, ROZEBOOM H J, WILLERS V P, et al.Discovery of a new class of bacterial heme-containing CC cleaving oxygenases[J].New Biotechnology, 2024, 83:82-90.
[13] TAVANTI M, HOSFORD J, LLOYD R C, et al.ATP regeneration by a single polyphosphate kinase powers multigram-scale aldehyde synthesis in vitro[J].Green Chemistry, 2021, 23(2):828-837.
[14] XIE X, WU P, HUANG X C, et al.Retro-protein XXA is a remarkable solubilizing fusion tag for inclusion bodies[J].Microbial Cell Factories, 2022, 21(1):51.
[15] YE X, LING X, WU M, et al.Improving soluble expression of SARS-CoV-2 spike priming protease TMPRSS2 with an artificial fusing protein[J].International Journal of Molecular Sciences, 2023, 24(13):10475.
[16] 敖龙, 朱玲玉, 庞欣, 等.人工蛋白XXA促进人源肿瘤抑素Tumstatin可溶性表达[J].食品与发酵工业, 2024, 50(12):1-8.
AO L, ZHU L Y, PANG X, et al.Artificial protein XXA promotes soluble expression of human-derived Tumstatin[J].Food and Fermentation Industries, 2024, 50(12):1-8.
[17] HEBDITCH M, CARBALLO-AMADOR M A, CHARONIS S, et al.Protein-Sol:A web tool for predicting protein solubility from sequence[J].Bioinformatics, 2017, 33(19):3098-3100.
[18] 邹树平, 颜海蔚, 胡忠策, 等.固定化重组大肠杆菌细胞催化合成(R)-环氧氯丙烷[J].现代化工, 2013, 33(7):55-59.
ZOU S P, YAN H W, HU Z C, et al.Synthesis of (R)-epichlorohydrin catalyzed by immobilized recombinant Escherichia coli cells[J].Mordern Chemical Industry, 2013, 33(7):55-59.
[19] 魏小平, 李建平, 蓝小月.分光光度法测定微量茴香醛的研究[J].食品科学, 2002, 23(1):101-103.
WEI X P, LI J P, LAN X Y.Spectrophotometric determination of micro-amount of anisaldehyde by condensation reaction with tba[J].Food Science, 2002, 23(1):101-103.
[20] 李沂馨, 毛银, 刘伟, 等.生物基1,6-己二醇的全细胞催化及其过程强化[J].食品与发酵工业, 2025, 51(5):8-14.
LI Y X, MAO Y, LIU W, et al.Whole-cell catalysis of biobased 1,6-hexanediol and its fermentation enhancement[J].Food and Fermentation Industries, 2025, 51(5):8-14.
[21] 付声亮, 邹诗瑶, 高娃, 等.重组大肠杆菌全细胞转化苹果酸合成丙酮酸[J].中国食品学报, 2024, 24(7):219-228.
FU S L, ZOU S Y, GAO W, et al.Biosynthesis of pyruvate from malic acid by whole-cell biotransformation using recombinant Escherichia coli[J].Journal of Chinese Institute of Food Science and Technology, 2024, 24(7):219-228.
[22] AUEZOVA L, NAJJAR A, KFOURY M, et al.Antibacterial activity of free or encapsulated selected phenylpropanoids against Escherichia coli and Staphylococcus epidermidis[J].Journal of Applied Microbiology, 2020, 128(3):710-720.
[23] LIN Y, HUANG R, SUN X X, et al.The p-Anisaldehyde/β-cyclodextrin inclusion complexes as a sustained release agent:Characterization, storage stability, antibacterial and antioxidant activity[J].Food Control, 2022, 132:108561.
[24] TAZON A W, AWWAD F, MEDDEB-MOUELHI F, et al.Biotechnological advances in vanillin production:From natural Vanilla to metabolic engineering platforms[J].BioChem, 2024, 4(4):323-349.
Options
文章导航

/

〈 〉

技术支持：北京玛格泰克科技发展有限公司