Heterologous expression of Fusarium oxysporum tomatinase in Saccharomyces cerevisiae increases its resistance to saponins and improves ethanol production during the fermentation of Agave tequilana Weber var. azul and Agave salmiana must |
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Authors: | Cira Luis Alberto González Gloria Angélica Torres Juan Carlos Pelayo Carlos Gutiérrez Melesio Ramírez Jesús |
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Institution: | (1) Centro de Investigación y Asistenta Tecnológica del Estado de Jalisco (CIATEJ), Unidad de Biotecnología, Av. Normalistas 800, Col. Colinas de la Normal, C.P. 44270, Guadalajara, Jalisco, Mexico;(2) Instituto de Investigación en Biología Experimental, Universidad de Guanajuato, Guanajuato, Mexico;(3) Centro Universitario de Ciencias exactas e Ingeniería de la Universidad de Guadalajara, Av. Marcelino García Barragán y Calzada Olímpica S.R., C.P. 44420, Guadalajara, Jalisco, Mexico |
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Abstract: | This paper describes the effect of the heterologous expression of tomatinase from Fusarium oxysporum f. sp lycopersici in Saccharomyces cerevisiae. The gene FoTom1 under the control of the S. cerevisiae phosphoglycerate kinase (PGK1) promoter was cloned into pYES2. S. cerevisiae strain Y45 was transformed with this vector and URA3 transformant strains were selected for resistance to α-tomatine. Two
transformants were randomly selected for further study (designated Y45-1 and Y45-2). Control strain Y45 was inhibited at 50 μM
α-tomatine, in contrast, transformants Y45-1 and Y45-2 did not show inhibition at 200 μM. Tomatinase activity was detected
by HPLC monitoring tomatine disappearance and tomatidine appearance in the supernatants of culture medium. Maximum tomatinase
activity was observed in the transformants after 6 h, remaining constant during the following 24 h. No tomatinase activity
was detected in the parental strain. Moreover, the transformants were able to grow and produce ethanol in a mix of Agave
tequilana Weber var. azul and Agave salmiana must, contrary to the Y45 strain which was unable to grow and ferment under these conditions. |
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Keywords: | Agave salmiana Agave tequilana Weber var azul Fusarium oxysporum Saccharomyces cerevisiae Saponins Tomatinase |
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