Iterative carotenogenic screens identify combinations of yeast gene deletions that enhance sclareol production

Microbial Cell Factories

Research

Codruta Ignea¹  Sotirios C Kampranis¹  Konstantia Kotta²  Aggeliki Andreadelli²  Antonios M Makris²  Anagnostis Argiriou²  Alexandros Nikolaidis²  Fotini A Trikka²  Anastasia Athanasakoglou³ 

[1] Department of Biochemistry, School of Medicine, University of Crete, P.O. Box 2208, 71003, Heraklion, Greece;Institute of Applied Biosciences/CERTH, P.O. Box 60361, Thermi, 57001, Thessaloniki, Greece;Institute of Applied Biosciences/CERTH, P.O. Box 60361, Thermi, 57001, Thessaloniki, Greece;Department of Biochemistry, School of Medicine, University of Crete, P.O. Box 2208, 71003, Heraklion, Greece;

关键词: Terpenoids;    Saccharomyces cerevisiae;    Sclareol;    Carotenoid;    Ergosterol biosynthesis;   

DOI  :  10.1186/s12934-015-0246-0

 received in 2015-02-04, accepted in 2015-04-17,  发布年份 2015

来源: Springer

【 摘 要 】

BackgroundTerpenoids (isoprenoids) have numerous applications in flavors, fragrances, drugs and biofuels. The number of microbially produced terpenoids is increasing as new biosynthetic pathways are being elucidated. However, efforts to improve terpenoid production in yeast have mostly taken advantage of existing knowledge of the sterol biosynthetic pathway, while many additional factors may affect the output of the engineered system.ResultsAiming to develop a yeast strain that can support high titers of sclareol, a diterpene of great importance for the perfume industry, we sought to identify gene deletions that improved carotenoid, and thus potentially sclareol, production. Using a carotenogenic screen, the best 100 deletion mutants, out of 4,700 mutant strains, were selected to create a subset for further analysis. To identify combinations of deletions that cooperate to further boost production, iterative carotenogenic screens were applied, and each time the top performing gene deletions were further ranked according to the number of genetic and physical interactions known for each specific gene. The gene selected in each round was deleted and the resulting strain was employed in a new round of selection. This approach led to the development of an EG60 derived haploid strain combining six deletions (rox1, dos2, yer134c, vba5, ynr063w and ygr259c) and exhibiting a 40-fold increase in carotenoid and 12-fold increase in sclareol titers, reaching 750 mg/L sclareol in shake flask cultivation.ConclusionUsing an iterative approach, we identified novel combinations of yeast gene deletions that improve carotenoid and sclareol production titers without compromising strain growth and viability. Most of the identified deletions have not previously been implicated in sterol pathway control. Applying the same approach using a different starting point could yield alternative sets of deletions with similar or improved outcome.

【 授权许可】

Unknown   
© Trikka et al.; licensee BioMed Central. 2015. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.