GIULIETTI Ana Maria
congresos y reuniones científicas
Cloning and yeast expression of Hyoscyamine 6Ò-hydroxylase as a tool for the development of an industrial biocatalyst..
CARDILLO, A; GIULIETTI, A M
Congreso; Biolatina 2006; 2006
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 (6b-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-b hydroxylase, the enzyme responsible for hyoscyamine hydroxylation and epoxidation, leading to scopolamine was isolated from Brugmansia candida. The h6hcDNA was cloned into pYES2.1 and pYES2.1-TOPO TA vectors to produce an untagged and tagged protein, respectively. The H6H enzyme was expressed 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 untagged proteins were able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The strains obtained in this work are promising and potentially applicable for biocatalytic processes