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Heterologous expression of a thermophilic esterase in Kluyveromyces yeasts

  • Applied Microbial and Cell Physiology
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Abstract

In the present work, a thermophilic esterase from Thermus thermophilus HB27 was cloned into Kluyveromyces marxianus and into Kluyveromyces lactis using two different expression systems, yielding four recombinant strains. K. lactis showed the highest esterase expression levels (294 units per gram dry cell weight, with 65% of cell-bound enzyme) using an episomal system with the PGK promoter and terminator from Saccharomyces cerevisiae combined with the K. lactis k1 secretion signal. K. marxianus showed higher secretion efficiency of the heterologous esterase (56.9 units per gram dry cell weight, with 34% of cell-bound enzyme) than K. lactis. Hydrolytic activities for the heterologous esterases were maximum at pH values between 8.0 and 9.0 for both yeast species and at temperatures of 50 °C and 45 °C for K. marxianus and K. lactis, respectively. When compared to previously published data on this same esterase produced in the original host or in S. cerevisiae, our results indicate that Kluyveromyces yeasts can be considered good hosts for the heterologous secretion of thermophilic esterases, which have a potential application in biodiesel production or in resolving racemates.

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Acknowledgments

We would like to thank Dr. Nancy da Silva (University of California at Irvine) for kindly providing us the pNADFL11 plasmid, Dr. Hiroshi Fukuhara for providing the pSPGK1 plasmid, and Dr. Wésolowski-Louvel for kindly donating the K. lactis PM5-3C strain. SNR acknowledges grants received from Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP, São Paulo, Brazil) and from Coordenadoria de Aperfeiçoamento de Pessoal de Nível Superior (CAPES, Brasília, Brazil), which made possible a 1-year internship for the researcher at the Biochemistry and Molecular Biology Laboratory of Universidade da Coruña (Spain), where the DNA work was carried out. This work was financially supported by FAPESP and by Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq, Brasília, Brazil), and by Xunta de Galicia (Proyecto PGIDIT06REM38302PR, Spain). General support to the group of the UDC was funded by “Consolidación” program from C.E.O.U. Xunta de Galicia cofinanced by FEDER.

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  1. Saul Nitsche Rocha

    Present address: Universidade Positivo, Rua Pedro Viriato Parigot de Souza, 5300, CEP 81280-330, Curitiba, Paraná, Brasil

Authors and Affiliations

  1. Department of Chemical Engineering, University of São Paulo, C.P. 61548, 05424-970, São Paulo, SP, Brazil

    Saul Nitsche Rocha & Andreas Karoly Gombert

  2. School of Pharmaceutical Sciences, University of São Paulo, Av. Prof. Lineu Prestes, 580-bloco 16, 05508-900, São Paulo, SP, Brazil

    José Abrahão-Neto

  3. Departamento de Bioloxía Celular e Molecular, Facultade de Ciencias, Universidade da Coruña, Campus da Zapateira s/n, 15071-A, Coruña, Spain

    María Esperanza Cerdán & María Isabel González-Siso

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  1. Saul Nitsche Rocha
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  2. José Abrahão-Neto
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  3. María Esperanza Cerdán
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  4. Andreas Karoly Gombert
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  5. María Isabel González-Siso
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Correspondence to María Isabel González-Siso.

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Rocha, S.N., Abrahão-Neto, J., Cerdán, M.E. et al. Heterologous expression of a thermophilic esterase in Kluyveromyces yeasts. Appl Microbiol Biotechnol 89, 375–385 (2011). https://doi.org/10.1007/s00253-010-2869-8

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  • DOI: https://doi.org/10.1007/s00253-010-2869-8

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