Efficient Production of l-Ribose with a Recombinant Escherichia coli Biocatalyst▿
| Autor: | Ryan Woodyer, F. Michael Racine, Shama N. Khan, Badal C. Saha, Nathan Wymer |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2008 |
| Předmět: |
Glycerol
Polymers Ribose Coenzymes Gene Expression Dehydrogenase medicine.disease_cause Ribitol Applied Microbiology and Biotechnology chemistry.chemical_compound Chlorides medicine Escherichia coli Cloning Molecular Apium Ecology biology Apium graveolens biology.organism_classification Physiology and Biotechnology Enterobacteriaceae Recombinant Proteins chemistry Biochemistry Biocatalysis Zinc Compounds Food Science Biotechnology Sugar Alcohol Dehydrogenases |
| Popis: | Optically pure carbohydrates are important intermediates for the preparation of pharmaceutical, food, and agrochemical products (2, 4, 15, 22). In particular, these carbohydrates are increasingly important in biochemical research and in development of new pharmaceutical therapies since carbohydrates are involved in cellular recognition, signaling, extra- and intracellular targeting, and even the development of disease states (1, 2, 7, 22, 27). Access to consistent, optically pure, and inexpensive carbohydrate starting materials is critical to the continuation of this research. The unique NAD-dependent mannitol-1-dehydrogenase (MDH) from Apium graveolens (33-38, 44) acts on the 1 position of d-mannitol, producing d-mannose (Fig. (Fig.1),1), in contrast to the more common 2-mannitol dehydrogenase, which interconverts d-mannitol and d-fructose (37). This novel regioselectivity combined with stringent stereoselectivity at the 2 position allows the MDH enzyme from A. graveolens to catalyze several interesting conversions, including the conversion of ribitol to l-ribose (Fig. (Fig.1),1), the conversion of d-sorbitol to l-gulose, and the conversion of galactitol to l-galactose (33). Sugars with the l configuration are often available only in limited amounts or at a high cost. To address these availability and economic concerns, utilization of MDH from A. graveolens is proposed. FIG. 1. MDH-catalyzed reactions. The MDH from A. graveolens catalyzes the unique conversion of d-mannitol to d-mannose and, similarly, the conversion of ribitol to l-ribose. The 2R stereochemistry is conserved for all of the MDH-catalyzed conversions. For this study, l-ribose was chosen as a model target since it is the potential starting material for many l-nucleoside-based pharmaceutical compounds, including Clevudine, Tyzeka, Valtorcitabine, Elvucitabine, and Epivir (12, 14, 40). The interest in l-nucleosides has increased, as noted in Table Table1,1, which shows several l-nucleoside-based pharmaceutical compounds recently approved or presently in clinical trials, creating a demand for l-ribose. Although several methods for the production of l-ribose have been described (19, 21, 31, 39), no method has provided an efficient and inexpensive source of l-ribose, as apparent from the increase in the bulk price (estimated at $1,000/kg, a price too high for intermediate-stage development) (29). With l-ribose selected as the model target for this platform technology, we created a novel recombinant Escherichia coli strain using a gene encoding the A. graveolens MDH. The recombinant MDH was characterized, and a process for whole-cell conversion was developed and improved, followed by liter scale production and isolation of l-ribose. TABLE 1. Examples of l-nucleoside-based pharmaceuticals |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|