Metabolic engineering of Saccharomyces cerevisiae for chelerythrine biosynthesis

被引:6
作者
Zhu, Jiawei [1 ]
Zhang, Kai [1 ]
He, Yuanzhi [1 ]
Zhang, Qi [1 ]
Ran, Yanpeng [1 ]
Tan, Zaigao [1 ]
Cui, Li [1 ]
Feng, Yan [1 ]
机构
[1] Shanghai Jiao Tong Univ, Sch Life Sci & Biotechnol, State Key Lab Microbial Metab, 800 Dongchuan Rd, Shanghai 200240, Peoples R China
基金
中国国家自然科学基金;
关键词
Chelerythrine; Metabolic engineering; Synthetic biology; Natural product biosynthesis; Saccharomyces cerevisiae; Microbial cell factory; BERBERINE BRIDGE ENZYME; BENZYLISOQUINOLINE ALKALOIDS; MOLECULAR-CLONING; YEAST; SANGUINARINE; EXPRESSION; PROVIDES; GENOME; POPPY; ACID;
D O I
10.1186/s12934-024-02448-4
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
Background Chelerythrine is an important alkaloid used in agriculture and medicine. However, its structural complexity and low abundance in nature hampers either bulk chemical synthesis or extraction from plants. Here, we reconstructed and optimized the complete biosynthesis pathway for chelerythrine from (S)-reticuline in Saccharomyces cerevisiae using genetic reprogramming. Results The first-generation strain Z4 capable of producing chelerythrine was obtained via heterologous expression of seven plant-derived enzymes (McoBBE, TfSMT, AmTDC, EcTNMT, PsMSH, EcP6H, and PsCPR) in S. cerevisiae W303-1 A. When this strain was cultured in the synthetic complete (SC) medium supplemented with 100 mu M of (S)-reticuline for 10 days, it produced up to 0.34 mu g/L chelerythrine. Furthermore, efficient metabolic engineering was performed by integrating multiple-copy rate-limiting genes (TfSMT, AmTDC, EcTNMT, PsMSH, EcP6H, PsCPR, INO2, and AtATR1), tailoring the heme and NADPH engineering, and engineering product trafficking by heterologous expression of MtABCG10 to enhance the metabolic flux of chelerythrine biosynthesis, leading to a nearly 900-fold increase in chelerythrine production. Combined with the cultivation process, chelerythrine was obtained at a titer of 12.61 mg per liter in a 0.5 L bioreactor, which is over 37,000-fold higher than that of the first-generation recombinant strain. Conclusions This is the first heterologous reconstruction of the plant-derived pathway to produce chelerythrine in a yeast cell factory. Applying a combinatorial engineering strategy has significantly improved the chelerythrine yield in yeast and is a promising approach for synthesizing functional products using a microbial cell factory. This achievement underscores the potential of metabolic engineering and synthetic biology in revolutionizing natural product biosynthesis.
引用
收藏
页数:13
相关论文
共 49 条
[1]   Safety and efficacy of a feed additive consisting of Macleaya cordata (Willd.) R. Br. extract and leaves (Sangrovit® extra) for all poultry species (excluding laying and breeding birds) (Phytobiotics Futterzusatzstoffe GmbH) [J].
Bampidis, Vasileios ;
Azimonti, Giovanna ;
Bastos, Maria de Lourdes ;
Christensen, Henrik ;
Dusemund, Birgit ;
Durjava, Mojca ;
Kouba, Maryline ;
Lopez-Alonso, Marta ;
Puente, Secundino Lopez ;
Marcon, Francesca ;
Mayo, Baltasar ;
Pechova, Alena ;
Petkova, Mariana ;
Ramos, Fernando ;
Sanz, Yolanda ;
Villa, Roberto Edoardo ;
Woutersen, Ruud ;
Brantom, Paul ;
Chesson, Andrew ;
Westendorf, Johannes ;
Manini, Paola ;
Casanova, Jordi Ortuno ;
Dusemund, Birgit .
EFSA JOURNAL, 2023, 21 (06)
[2]   Isolation and characterization of a cDNA encoding (S)-cis-N-methylstylopine 14-hydroxylase from opium poppy, a key enzyme in sanguinarine biosynthesis [J].
Beaudoin, Guillaume A. W. ;
Facchini, Peter J. .
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 2013, 431 (03) :597-603
[3]   Berberine bridge enzyme, a key branch-point enzyme in benzylisoquinoline alkaloid biosynthesis, contains a vacuolar sorting determinant [J].
Bird, DA ;
Facchini, PJ .
PLANTA, 2001, 213 (06) :888-897
[4]   Naturally-Occurring Alkaloids of Plant Origin as Potential Antimicrobials against Antibiotic-Resistant Infections [J].
Casciaro, Bruno ;
Mangiardi, Laura ;
Cappiello, Floriana ;
Romeo, Isabella ;
Loffredo, Maria Rosa ;
Iazzetti, Antonia ;
Calcaterra, Andrea ;
Goggiamani, Antonella ;
Ghirga, Francesca ;
Mangoni, Maria Luisa ;
Botta, Bruno ;
Quaglio, Deborah .
MOLECULES, 2020, 25 (16)
[5]   History of the use of antibiotic as growth promoters in European poultry feeds [J].
Castanon, J. I. R. .
POULTRY SCIENCE, 2007, 86 (11) :2466-2471
[6]   Characterization of two methylenedioxy bridge-forming cytochrome P450-dependent enzymes of alkaloid formation in the Mexican prickly poppy Argemone mexicana [J].
Chavez, Maria Luisa Diaz ;
Rolf, Megan ;
Gesell, Andreas ;
Kutchan, Toni M. .
ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS, 2011, 507 (01) :186-193
[7]   Engineering cofactor supply and recycling to drive phenolic acid biosynthesis in yeast [J].
Chen, Ruibing ;
Gao, Jiaoqi ;
Yu, Wei ;
Chen, Xianghui ;
Zhai, Xiaoxin ;
Chen, Yu ;
Zhang, Lei ;
Zhou, Yongjin J. .
NATURE CHEMICAL BIOLOGY, 2022, 18 (05) :520-+
[8]   Synthetic biology strategies for microbial biosynthesis of plant natural products [J].
Cravens, Aaron ;
Payne, James ;
Smolke, Christina D. .
NATURE COMMUNICATIONS, 2019, 10 (1)
[9]   An enzyme-coupled biosensor enables (S)-reticuline production in yeast from glucose [J].
DeLoache, William C. ;
Russ, Zachary N. ;
Narcross, Lauren ;
Gonzales, Andrew M. ;
Martin, Vincent J. J. ;
Dueber, John E. .
NATURE CHEMICAL BIOLOGY, 2015, 11 (07) :465-+
[10]   MOLECULAR-CLONING, EXPRESSION, AND INDUCTION OF BERBERINE BRIDGE ENZYME, AN ENZYME ESSENTIAL TO THE FORMATION OF BENZOPHENANTHRIDINE ALKALOIDS IN THE RESPONSE OF PLANTS TO PATHOGENIC ATTACK [J].
DITTRICH, H ;
KUTCHAN, TM .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1991, 88 (22) :9969-9973