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Efficient secretion of Bacillus subtilis lipase A in Saccharomyces cerevisiae by translational fusion to the Pir4 cell wall protein

  • Applied Genetics and Molecular Biotechnology
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Abstract

Both the secretion and the cell surface display of Bacillus subtilis lipase A (Lip A) in Saccharomyces cerevisiae was investigated using different domains of the cell wall protein Pir4 as translational fusion partners. LipA gene minus its leader peptide was fused inframe in two places of PIR4 to achieve cell wall targeting, or substituting most of the PIR4 sequence, after the signal peptide and the Kex2 processed subunit I of Pir4 to achieve secretion to the growth medium. Expression of the recombinant fusion proteins was investigated in a standard and a glycosylation-deficient strain of S. cerevisiae, grown in selective or rich medium. Fusion proteins intended to be retained at the cell wall were secreted to the growth medium, most likely as result of the degradation of the Pir4 moiety containing the cell wall retention domain, giving low levels of lipase activity. However, the fusion intended for secretion was efficiently secreted in a percentage of close to 90% and remained stable even in rich medium at high cell density cultures, yielding values of over 400 IU of lipase activity per milliliter of cell supernatant. This is, to our knowledge, the first report of the efficient production, as a secreted protein, of lipase A of B. subtilis in baker’s yeast.

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Acknowledgements

This work was supported by grant ISCIII2006-PI0731 from the Ministerio de Sanidad/Instituto de la Salud Carlos III (Spain). María Mormeneo and Cristina Bofill were recipient of predoctoral grants from the Programa Nacional de Formación de Profesorado Universitario del Ministerio de Educación y Ciencia. Isabel Andrés was the recipient of a predoctoral grant from the Generalitat Valenciana.

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Authors and Affiliations

  1. Unidad de Microbiología. Facultad de Farmacia, Univ. De Valencia. Avda, Vicente Andrés Estelles s/n, 46100, Burjassot, Valencia, Spain

    María Mormeneo, Isabel Andrés & Jesús Zueco

  2. Department Microbiología. Fac. Biología, Univ. Barcelona, Avenida Diagonal 645, 08028, Barcelona, Spain

    Cristina Bofill & Pilar Díaz

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  1. María Mormeneo
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  2. Isabel Andrés
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  3. Cristina Bofill
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  4. Pilar Díaz
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  5. Jesús Zueco
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Correspondence to Jesús Zueco.

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Mormeneo, M., Andrés, I., Bofill, C. et al. Efficient secretion of Bacillus subtilis lipase A in Saccharomyces cerevisiae by translational fusion to the Pir4 cell wall protein. Appl Microbiol Biotechnol 80, 437–445 (2008). https://doi.org/10.1007/s00253-008-1549-4

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  • DOI: https://doi.org/10.1007/s00253-008-1549-4

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