High-yield ‘one-pot’ biosynthesis of raspberry ketone, a high-value fine chemical
| Autor: | Paul S. Freemont, Tommaso Tosi, Simon J. Moore, Derek Bell, Yonek B Hleba, Karen M. Polizzi |
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| Přispěvatelé: | Engineering & Physical Science Research Council (EPSRC) |
| Rok vydání: | 2021 |
| Předmět: |
AcademicSubjects/SCI01060
AcademicSubjects/SCI02299 AcademicSubjects/SCI02298 Biomedical Engineering Raspberry ketone Bioengineering Reductase fine chemical Cofactor Biomaterials QH301 chemistry.chemical_compound Biosynthesis Organic chemistry cell-free biology Artificial enzyme Agricultural and Biological Sciences (miscellaneous) QR Blowing a raspberry chemistry QD431 raspberry ketone Yield (chemistry) biology.protein AcademicSubjects/SCI00520 Fine chemical AcademicSubjects/SCI00980 synthetic biology Research Article polyketides Biotechnology |
| Zdroj: | Synthetic Biology |
| ISSN: | 2397-7000 |
| Popis: | Cell-free extract and purified enzyme-based systems provide an attractive solution to study biosynthetic strategies towards a range of chemicals. 4-(4-hydroxyphenyl)-butan-2-one, also known as raspberry ketone, is the major fragrance component of raspberry fruit and is used as a natural additive in the food and sports industry. Current industrial processing of the natural form of raspberry ketone involves chemical extraction from a yield of ∼1–4 mg kg−1 of fruit. Due to toxicity, microbial production provides only low yields of up to 5–100 mg L−1. Herein, we report an efficient cell-free strategy to probe into a synthetic enzyme pathway that converts either L-tyrosine or the precursor, 4-(4-hydroxyphenyl)-buten-2-one, into raspberry ketone at up to 100% conversion. As part of this strategy, it is essential to recycle inexpensive cofactors. Specifically, the final enzyme step in the pathway is catalyzed by raspberry ketone/zingerone synthase (RZS1), an NADPH-dependent double bond reductase. To relax cofactor specificity towards NADH, the preferred cofactor for cell-free biosynthesis, we identify a variant (G191D) with strong activity with NADH. We implement the RZS1 G191D variant within a ‘one-pot’ cell-free reaction to produce raspberry ketone at high-yield (61 mg L−1), which provides an alternative route to traditional microbial production. In conclusion, our cell-free strategy complements the growing interest in engineering synthetic enzyme cascades towards industrially relevant value-added chemicals. |
| Databáze: | OpenAIRE |
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