Expression of Brugmansia candida Hyoscyamine 6beta-Hydroxylase gene in Saccharomyces cerevisiae and its potential use as biocatalyst
Open Access
- 27 May 2008
- journal article
- Published by Springer Science and Business Media LLC in Microbial Cell Factories
- Vol. 7 (1), 17
- https://doi.org/10.1186/1475-2859-7-17
Abstract
Tropane alkaloids, mainly hyoscyamine and scopolamine, are widely used in medicine due to their anticholinergic activity. Scopolamine has a higher demand being the more valuable alkaloid due to its fewer side effects and higher physiological activity. Anisodamine (6β-hydroxyhyoscyamine) is the intermediate in the conversion of hyoscyamine into scopolamine. Current studies report that this alkaloid is potentially applicable in medicine. The gene that codifies for Hyoscyamine 6-β hydroxylase, the enzyme responsible for hyoscyamine hydroxylation and epoxidation, leading to scopolamine was isolated from Brugmansia candida. The h6h cDNA was cloned into pYES2.1 and pYES2.1/V5-His-TOPO vectors to produce an untagged and a tagged protein, respectively. The H6H enzyme was produced in Saccharomyces cerevisiae in order to obtain a biological catalyst for potential industrial applications. Protein extracts of the induced yeast were analyzed by Western blot. The expression was detected 4 h after induction and no degradation was observed during the period assayed. The tagged and the untagged proteins were able to transform hyoscyamine, showing a functional expression of the h6h cDNA. The strains obtained in this work are promising and potentially applicable in biocatalytic processes.Keywords
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