Tailored Biosynthesis of Gibberellin Plant Hormones in Yeast

Overview

Journal Metab Eng

Specialties Biomedical Engineering
Endocrinology

Date 2021 Mar 22

PMID 33746070

Citations 19

Authors

Kanchana R Kildegaard

Jonathan A Arnesen

Belen Adiego-Perez

Daniela Rago

Mette Kristensen

Andreas K Klitgaard

Esben H Hansen

Jorgen Hansen

Irina Borodina

Affiliations

Soon will be listed here.

Abstract

The application of small amounts of natural plant growth hormones, such as gibberellins (GAs), can increase the productivity and quality of many vegetable and fruit crops. However, gibberellin growth hormones usage is limited by the high cost of their production, which is currently based on fermentation of a natural fungal producer Fusarium fujikuroi that produces a mix of several GAs. We explored the potential of the oleaginous yeast Yarrowia lipolytica to produce specific profiles of GAs. Firstly, the production of the GA-precursor ent-kaurenoic acid (KA) at 3.75 mg/L was achieved by expression of biosynthetic enzymes from the plant Arabidopsis thaliana and upregulation of the mevalonate (MVA) pathway. We then built a GA-producing strain by extending the GA-biosynthetic pathway and upregulating the MVA-pathway further, resulting in 17.29 mg/L GA. Additional expression of the F. fujikoroi GA-biosynthetic enzymes resulted in the production of GA (trace amounts) and GA (2.93 mg/L). Lastly, through protein engineering and the expression of additional KA-biosynthetic genes, we increased the GA-production 4.4-fold resulting in 12.81 mg/L. The developed system presents a promising resource for the recombinant production of specific gibberellins, identifying bottlenecks in GA biosynthesis, and discovering new GA biosynthetic genes. CLASSIFICATION: Biological Sciences, Applied Biological Sciences.

Citing Articles

Optimizing longifolene production in via metabolic and protein engineering.

Yao Y, Zhang X, Wang C, Zhang Y, Li D, Yang W Synth Syst Biotechnol. 2025; 10(2):433-441.

PMID: 39925943 PMC: 11803839. DOI: 10.1016/j.synbio.2025.01.004.

Sustainable biosynthesis of valuable diterpenes in microbes.

Liu Y, Chen X, Zhang C Eng Microbiol. 2024; 3(1):100058.

PMID: 39628524 PMC: 11611012. DOI: 10.1016/j.engmic.2022.100058.

Differential symbiotic compatibilities between rhizobium strains and cultivated and wild soybeans revealed by anatomical and transcriptome analyses.

Zadegan S, Kim W, Abbas H, Kim S, Krishnan H, Hewezi T Front Plant Sci. 2024; 15:1435632.

PMID: 39290740 PMC: 11405202. DOI: 10.3389/fpls.2024.1435632.

Strangers in a foreign land: 'Yeastizing' plant enzymes.

Van Gelder K, Lindner S, Hanson A, Zhou J Microb Biotechnol. 2024; 17(9):e14525.

PMID: 39222378 PMC: 11368087. DOI: 10.1111/1751-7915.14525.

Advances in the Structures, Pharmacological Activities, and Biosynthesis of Plant Diterpenoids.

Li L, Fu J, Liu N J Microbiol Biotechnol. 2024; 34(8):1563-1579.

PMID: 39081244 PMC: 11380518. DOI: 10.4014/jmb.2402.02014.

References

Darvishi F, Ariana M, Marella E, Borodina I . Advances in synthetic biology of oleaginous yeast Yarrowia lipolytica for producing non-native chemicals. Appl Microbiol Biotechnol. 2018; 102(14):5925-5938. DOI: 10.1007/s00253-018-9099-x. View

Lange T, Hedden P, Graebe J . Expression cloning of a gibberellin 20-oxidase, a multifunctional enzyme involved in gibberellin biosynthesis. Proc Natl Acad Sci U S A. 1994; 91(18):8552-6. PMC: 44644. DOI: 10.1073/pnas.91.18.8552. View

Prisic S, Peters R . Synergistic substrate inhibition of ent-copalyl diphosphate synthase: a potential feed-forward inhibition mechanism limiting gibberellin metabolism. Plant Physiol. 2007; 144(1):445-54. PMC: 1913771. DOI: 10.1104/pp.106.095208. View

Shi T, Peng H, Zeng S, Ji R, Shi K, Huang H . Microbial production of plant hormones: Opportunities and challenges. Bioengineered. 2016; 8(2):124-128. PMC: 5398602. DOI: 10.1080/21655979.2016.1212138. View

Holkenbrink C, Dam M, Kildegaard K, Beder J, Dahlin J, Domenech Belda D . EasyCloneYALI: CRISPR/Cas9-Based Synthetic Toolbox for Engineering of the Yeast Yarrowia lipolytica. Biotechnol J. 2018; 13(9):e1700543. DOI: 10.1002/biot.201700543. View

Escamilla E, Dendooven L, Magana I, Parra R, de la Torre M . Optimization of gibberellic acid production by immobilized Gibberella fujikuroi mycelium in fluidized bioreactors. J Biotechnol. 2000; 76(2-3):147-55. DOI: 10.1016/s0168-1656(99)00182-0. View

Rademacher W, Graebe J . Gibberellin A4 produced by Sphaceloma manihoticola, the cause of the superelongation disease of cassava (Manihot esculenta). Biochem Biophys Res Commun. 1979; 91(1):35-40. DOI: 10.1016/0006-291x(79)90579-5. View

Donald K, Hampton R, Fritz I . Effects of overproduction of the catalytic domain of 3-hydroxy-3-methylglutaryl coenzyme A reductase on squalene synthesis in Saccharomyces cerevisiae. Appl Environ Microbiol. 1997; 63(9):3341-4. PMC: 168639. DOI: 10.1128/aem.63.9.3341-3344.1997. View

Bottini R, Fulchieri M, Pearce D, Pharis R . Identification of Gibberellins A(1), A(3), and Iso-A(3) in Cultures of Azospirillum lipoferum. Plant Physiol. 1989; 90(1):45-7. PMC: 1061674. DOI: 10.1104/pp.90.1.45. View

10.

Gao S, Tong Y, Zhu L, Ge M, Zhang Y, Chen D . Iterative integration of multiple-copy pathway genes in Yarrowia lipolytica for heterologous β-carotene production. Metab Eng. 2017; 41:192-201. DOI: 10.1016/j.ymben.2017.04.004. View

11.

Hedden P, Sponsel V . A Century of Gibberellin Research. J Plant Growth Regul. 2015; 34(4):740-60. PMC: 4622167. DOI: 10.1007/s00344-015-9546-1. View

12.

Tudzynski B, Mihlan M, Rojas M, Linnemannstons P, Gaskin P, Hedden P . Characterization of the final two genes of the gibberellin biosynthesis gene cluster of Gibberella fujikuroi: des and P450-3 encode GA4 desaturase and the 13-hydroxylase, respectively. J Biol Chem. 2003; 278(31):28635-43. DOI: 10.1074/jbc.M301927200. View

13.

Durairaj P, Jung E, Park H, Kim B, Yun H . Comparative functional characterization of a novel benzoate hydroxylase cytochrome P450 of Fusarium oxysporum. Enzyme Microb Technol. 2015; 70:58-65. DOI: 10.1016/j.enzmictec.2014.12.013. View

14.

Hayashi K, Kawaide H, Notomi M, Sakigi Y, Matsuo A, Nozaki H . Identification and functional analysis of bifunctional ent-kaurene synthase from the moss Physcomitrella patens. FEBS Lett. 2006; 580(26):6175-81. DOI: 10.1016/j.febslet.2006.10.018. View

15.

Hamayun M, Khan S, Khan A, Shin J, Ahmad B, Shin D . Exogenous gibberellic acid reprograms soybean to higher growth and salt stress tolerance. J Agric Food Chem. 2010; 58(12):7226-32. DOI: 10.1021/jf101221t. View

16.

Kasahara H, Hanada A, Kuzuyama T, Takagi M, Kamiya Y, Yamaguchi S . Contribution of the mevalonate and methylerythritol phosphate pathways to the biosynthesis of gibberellins in Arabidopsis. J Biol Chem. 2002; 277(47):45188-94. DOI: 10.1074/jbc.M208659200. View

17.

Cao X, Lv Y, Chen J, Imanaka T, Wei L, Hua Q . Metabolic engineering of oleaginous yeast for limonene overproduction. Biotechnol Biofuels. 2016; 9:214. PMC: 5057495. DOI: 10.1186/s13068-016-0626-7. View

18.

Helliwell C, Sullivan J, Mould R, Gray J, Peacock W, Dennis E . A plastid envelope location of Arabidopsis ent-kaurene oxidase links the plastid and endoplasmic reticulum steps of the gibberellin biosynthesis pathway. Plant J. 2001; 28(2):201-8. DOI: 10.1046/j.1365-313x.2001.01150.x. View

19.

Kawaide H, Hayashi K, Kawanabe R, Sakigi Y, Matsuo A, Natsume M . Identification of the single amino acid involved in quenching the ent-kauranyl cation by a water molecule in ent-kaurene synthase of Physcomitrella patens. FEBS J. 2010; 278(1):123-33. DOI: 10.1111/j.1742-4658.2010.07938.x. View

20.

Tudzynski B, Kawaide H, Kamiya Y . Gibberellin biosynthesis in Gibberella fujikuroi: cloning and characterization of the copalyl diphosphate synthase gene. Curr Genet. 1998; 34(3):234-40. DOI: 10.1007/s002940050392. View