Hyoscyamine 6-â-hydroxylase, a recombinant biocatalyst for the industrial production of scopolamine

CARDILLO AB; RODRÍGUEZ TALOU J; GIULIETTI AM

Dalian, China

Simposio; 13th International Biotechnology Symposium and Meeting; 2008

International Biotechnology Symposium

The tropane alkaloids, hyoscyamine and scopolamine are widely used as pharmaceuticals due to their anticholinergic activity. Scopolamine is the more valuable, with a demand 10 times higher than that of hyoscyamine (Palazón et al., 2003; Hashimoto and Yamada, 1986). Brugmansia candida is a South American native plant and a tropane alkaloid producer. The conversion of hyoscyamine into scopolamine is carried out by hyoscyamine 6-- hydroxylase (H6H, EC 1.14.11.11). The development of a recombinant cell harbouring the plant enzyme constitutes a useful strategy for performing pharmaceutical processes. This work reports the functional expression of H6H enzyme in Saccharomyces cerevisiae native plant and a tropane alkaloid producer. The conversion of hyoscyamine into scopolamine is carried out by hyoscyamine 6-- hydroxylase (H6H, EC 1.14.11.11). The development of a recombinant cell harbouring the plant enzyme constitutes a useful strategy for performing pharmaceutical processes. This work reports the functional expression of H6H enzyme in Saccharomyces cerevisiae and Yamada, 1986). Brugmansia candida is a South American native plant and a tropane alkaloid producer. The conversion of hyoscyamine into scopolamine is carried out by hyoscyamine 6-- hydroxylase (H6H, EC 1.14.11.11). The development of a recombinant cell harbouring the plant enzyme constitutes a useful strategy for performing pharmaceutical processes. This work reports the functional expression of H6H enzyme in Saccharomyces cerevisiae native plant and a tropane alkaloid producer. The conversion of hyoscyamine into scopolamine is carried out by hyoscyamine 6-- hydroxylase (H6H, EC 1.14.11.11). The development of a recombinant cell harbouring the plant enzyme constitutes a useful strategy for performing pharmaceutical processes. This work reports the functional expression of H6H enzyme in Saccharomyces cerevisiae Palazón et al., 2003; Hashimoto and Yamada, 1986). Brugmansia candida is a South American native plant and a tropane alkaloid producer. The conversion of hyoscyamine into scopolamine is carried out by hyoscyamine 6-- hydroxylase (H6H, EC 1.14.11.11). The development of a recombinant cell harbouring the plant enzyme constitutes a useful strategy for performing pharmaceutical processes. This work reports the functional expression of H6H enzyme in Saccharomyces cerevisiae native plant and a tropane alkaloid producer. The conversion of hyoscyamine into scopolamine is carried out by hyoscyamine 6-- hydroxylase (H6H, EC 1.14.11.11). The development of a recombinant cell harbouring the plant enzyme constitutes a useful strategy for performing pharmaceutical processes. This work reports the functional expression of H6H enzyme in Saccharomyces cerevisiae ). Brugmansia candida is a South American native plant and a tropane alkaloid producer. The conversion of hyoscyamine into scopolamine is carried out by hyoscyamine 6-- hydroxylase (H6H, EC 1.14.11.11). The development of a recombinant cell harbouring the plant enzyme constitutes a useful strategy for performing pharmaceutical processes. This work reports the functional expression of H6H enzyme in Saccharomyces cerevisiaeSaccharomyces cerevisiae as a potential tool for the industrial production of scopolamine. The gene that codifies for the H6H enzyme was amplified from B. candida immature anthers. The h6h cDNA obtained was cloned into the pYES2 and the pYES2.1-TOPO TA vectors to produce an untagged and a tagged enzyme, respectively. The constructions were introduced in S. cerevisiae CEN PK2. The recombinant yeast strains obtained were induced for the expression with galactose. It was seen that the expression of the recombinant protein starts 4 h after induction. Crude protein extracts of the induced strains were assayed for the enzyme activity (Hashimoto and Yamada, 1987; Liu et al., 2005). The activity assay was incubated at 30 ◦C for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. 1987; Liu et al., 2005). The activity assay was incubated at 30 ◦C for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. strains obtained were induced for the expression with galactose. It was seen that the expression of the recombinant protein starts 4 h after induction. Crude protein extracts of the induced strains were assayed for the enzyme activity (Hashimoto and Yamada, 1987; Liu et al., 2005). The activity assay was incubated at 30 ◦C for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. 1987; Liu et al., 2005). The activity assay was incubated at 30 ◦C for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. into the pYES2 and the pYES2.1-TOPO TA vectors to produce an untagged and a tagged enzyme, respectively. The constructions were introduced in S. cerevisiae CEN PK2. The recombinant yeast strains obtained were induced for the expression with galactose. It was seen that the expression of the recombinant protein starts 4 h after induction. Crude protein extracts of the induced strains were assayed for the enzyme activity (Hashimoto and Yamada, 1987; Liu et al., 2005). The activity assay was incubated at 30 ◦C for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. 1987; Liu et al., 2005). The activity assay was incubated at 30 ◦C for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. strains obtained were induced for the expression with galactose. It was seen that the expression of the recombinant protein starts 4 h after induction. Crude protein extracts of the induced strains were assayed for the enzyme activity (Hashimoto and Yamada, 1987; Liu et al., 2005). The activity assay was incubated at 30 ◦C for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. 1987; Liu et al., 2005). The activity assay was incubated at 30 ◦C for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. immature anthers. The h6h cDNA obtained was cloned into the pYES2 and the pYES2.1-TOPO TA vectors to produce an untagged and a tagged enzyme, respectively. The constructions were introduced in S. cerevisiae CEN PK2. The recombinant yeast strains obtained were induced for the expression with galactose. It was seen that the expression of the recombinant protein starts 4 h after induction. Crude protein extracts of the induced strains were assayed for the enzyme activity (Hashimoto and Yamada, 1987; Liu et al., 2005). The activity assay was incubated at 30 ◦C for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. 1987; Liu et al., 2005). The activity assay was incubated at 30 ◦C for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. strains obtained were induced for the expression with galactose. It was seen that the expression of the recombinant protein starts 4 h after induction. Crude protein extracts of the induced strains were assayed for the enzyme activity (Hashimoto and Yamada, 1987; Liu et al., 2005). The activity assay was incubated at 30 ◦C for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. 1987; Liu et al., 2005). The activity assay was incubated at 30 ◦C for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. S. cerevisiae CEN PK2. The recombinant yeast strains obtained were induced for the expression with galactose. It was seen that the expression of the recombinant protein starts 4 h after induction. Crude protein extracts of the induced strains were assayed for the enzyme activity (Hashimoto and Yamada, 1987; Liu et al., 2005). The activity assay was incubated at 30 ◦C for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. 1987; Liu et al., 2005). The activity assay was incubated at 30 ◦C for 15 h. The analysis of the alkaloids was carried out by HPLC. The mobile phase used was octanesulfonic acid 0.01M pH 3/methanol (65:35), flow rate 1 ml/min. The results showed that the tagged and untagged enzymeswere able to transform hyoscyamine, showing a functional expression of the h6hcDNA. The untagged enzyme presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation. are promissory for technological applications in the production of scopolamine by biotransformation. presented a higher rate of conversion of hyoscyamine than the tagged enzyme and was able to produce scopolamine and not only 6--hydroxyhyoscyamine in the incubation times assayed. It can be concluded that the recombinant S. cerevisiae strains obtained are promissory for technological applications in the production of scopolamine by biotransformation.