Biosynthesis of the Antibiotic Nonribosomal Peptide Penicillin in Baker’s Yeast

Awan, Ali R.; Blount, Benjamin A.; Bell, David; Ho, Jack C H; McKiernan, Robert M; Ellis, Tom

doi:10.1101/075325

Cited by 2 publications

(3 citation statements)

References 27 publications

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“…The strategy, summarized in Figure , consists in a digestion‐ligation reaction guided by BsaI defined sites where the three promoters can be introduced in each of the three promoter positions in the car‐cassette. It has been recently proven by sequencing the Golden Gate reaction products that every possible combination is unbiased generated during combinatorial assemblies (Awan et al, ). Then, the pool of cassettes were simultaneously amplified, linearized and used to transform the wild type parental strain (wt), where the difference in color intensity could be easily screened (Supplementary Figure S4).…”

Section: Resultsmentioning

confidence: 99%

A synthetic biology approach to transform Yarrowia lipolytica into a competitive biotechnological producer of β‐carotene

Larroude

Celińska

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et al. 2017

Biotech & Bioengineering

275

237

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The increasing market demands of β-carotene as colorant, antioxidant and vitamin precursor, requires novel biotechnological production platforms. Yarrowia lipolytica, is an industrial organism unable to naturally synthesize carotenoids but with the ability to produce high amounts of the precursor Acetyl-CoA. We first found that a lipid overproducer strain was capable of producing more β-carotene than a wild type after expressing the heterologous pathway. Thereafter, we developed a combinatorial synthetic biology approach base on Golden Gate DNA assembly to screen the optimum promoter-gene pairs for each transcriptional unit expressed. The best strain reached a production titer of 1.5 g/L and a maximum yield of 0.048 g/g of glucose in flask. β-carotene production was further increased in controlled conditions using a fed-batch fermentation. A total production of β-carotene of 6.5 g/L and 90 mg/g DCW with a concomitant production of 42.6 g/L of lipids was achieved. Such high titers suggest that engineered Y. lipolytica is a competitive producer organism of β-carotene.

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Section: Resultsmentioning

confidence: 99%

A synthetic biology approach to transform Yarrowia lipolytica into a competitive biotechnological producer of β‐carotene

Larroude

Celińska

Back

et al. 2017

Biotech & Bioengineering

275

237

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“…Firstly, we used a large-scale combinatorial approach to metabolic pathway construction. Such approaches have proven useful for simultaneously optimising expression of multiple genes in other organisms 37,38,45,46 , but have not been established in cyanobacteria. Only two small-scale examples of combinatorial designs have been reported, the first varying a small number of alternative enzyme-encoding sequences 60 and the second varying only the RBSs in a two-enzyme pathway 72 .…”

Section: Discussionmentioning

confidence: 99%

“…The same assembly methods allow for optimisation by rational replacement of individual parts to fine-tune expression. This combinatorial approach to construction and optimisation of metabolic pathway-encoding constructs is a recent development, and while its effectiveness has been clearly demonstrated in model organisms like Escherichia coli and Saccharomyces cerevisae 37,38,45,46 , it is not yet standard practice, and in many other organisms including cyanobacteria it has not yet been established.…”

Section: Introductionmentioning

confidence: 99%

Combinatorial metabolic engineering platform enabling stable overproduction of lycopene from carbon dioxide by cyanobacteria

Taylor

Heap

2020

Preprint

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Cyanobacteria are simple, efficient, genetically-tractable photosynthetic microorganisms representing ideal biocatalysts for CO 2 capture and conversion, in principle. In practice, genetic instability and low productivity are key, linked problems in engineered cyanobacteria. We took a massively parallel approach, generating and characterising libraries of synthetic promoters and RBSs for the cyanobacterium Synechocystis , and assembling a sparse combinatorial library of millions of metabolic pathway-encoding construct variants. Laboratory evolution suppressed variants causing metabolic burden in Synechocystis , leading to expected genetic instability. Surprisingly however, in a single combinatorial round without iterative optimisation, 80% of variants chosen at random overproduced the valuable terpenoid lycopene from atmospheric CO 2 over many generations, apparently overcoming the trade-off between stability and productivity. This first large-scale parallel metabolic engineering of cyanobacteria provides a new platform for development of genetically stable cyanobacterial biocatalysts for sustainable light-driven production of valuable products directly from CO 2 , avoiding fossil carbon or competition with food production.

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Biosynthesis of the Antibiotic Nonribosomal Peptide Penicillin in Baker’s Yeast

Cited by 2 publications

References 27 publications

A synthetic biology approach to transform Yarrowia lipolytica into a competitive biotechnological producer of β‐carotene

A synthetic biology approach to transform Yarrowia lipolytica into a competitive biotechnological producer of β‐carotene

Combinatorial metabolic engineering platform enabling stable overproduction of lycopene from carbon dioxide by cyanobacteria

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