caa a22 4500 605503125 CHVBK 20210128100609.0 cr unu---uuuuu 210128e20150701xx s 000 0 eng 10.1007/s00253-014-6328-9 doi (NATIONALLICENCE)springer-10.1007/s00253-014-6328-9 Strategies for the production of difficult-to-express full-length eukaryotic proteins using microbial cell factories: production of human alpha-galactosidase A [Elektronische Daten] [Ugutz Unzueta, Felicitas Vázquez, Giulia Accardi, Rosa Mendoza, Verónica Toledo-Rubio, Maria Giuliani, Filomena Sannino, Ermenegilda Parrilli, Ibane Abasolo, Simo Schwartz Jr., Maria Tutino, Antonio Villaverde, José Corchero, Neus Ferrer-Miralles] Obtaining high levels of pure proteins remains the main bottleneck of many scientific and biotechnological studies. Among all the available recombinant expression systems, Escherichia coli facilitates gene expression by its relative simplicity, inexpensive and fast cultivation, well-known genetics and the large number of tools available for its biotechnological application. However, recombinant expression in E. coli is not always a straightforward procedure and major obstacles are encountered when producing many eukaryotic proteins and especially membrane proteins, linked to missing posttranslational modifications, proteolysis and aggregation. In this context, many conventional and unconventional eukaryotic hosts are under exploration and development, but in some cases linked to complex culture media or processes. In this context, alternative bacterial systems able to overcome some of the limitations posed by E. coli keeping the simplicity of prokaryotic manipulation are currently emerging as convenient hosts for protein production. We have comparatively produced a "difficult-to-express” human protein, the lysosomal enzyme alpha-galactosidase A (hGLA) in E. coli and in the psychrophilic bacterium Pseudoalteromonas haloplanktis TAC125 cells (P. haloplanktis TAC125). While in E. coli the production of active hGLA was unreachable due to proteolytic instability and/or protein misfolding, the expression of hGLA gene in P. haloplanktis TAC125 allows obtaining active enzyme. These results are discussed in the context of emerging bacterial systems for protein production that represent appealing alternatives to the regular use of E. coli and also of more complex eukaryotic systems. Springer-Verlag Berlin Heidelberg, 2015 Recombinant protein nationallicence Expression systems nationallicence Escherichia coli nationallicence Pseudoalteromonas haloplanktis TAC125 nationallicence Human alpha-galactosidase A nationallicence Fabry's disease nationallicence Unzueta Ugutz Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut Vázquez Felicitas Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut Accardi Giulia Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut Mendoza Rosa Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut Toledo-Rubio Verónica Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut Giuliani Maria Dipartimento di Scienze Biochimiche, Facoltà di Medicina e Chirurgia, Università degli studi di Palermo, 90127, Palermo, Italy aut Sannino Filomena Dipartimento di Scienze Biochimiche, Facoltà di Medicina e Chirurgia, Università degli studi di Palermo, 90127, Palermo, Italy aut Parrilli Ermenegilda Dipartimento di Scienze Biochimiche, Facoltà di Medicina e Chirurgia, Università degli studi di Palermo, 90127, Palermo, Italy aut Abasolo Ibane CIBER de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Barcelona, Spain aut Schwartz Jr. Simo CIBER de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Barcelona, Spain aut Tutino Maria Dipartimento di Scienze Biochimiche, Facoltà di Medicina e Chirurgia, Università degli studi di Palermo, 90127, Palermo, Italy aut Villaverde Antonio Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut Corchero José Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut Ferrer-Miralles Neus Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/14(2015-07-01), 5863-5874 0175-7598 99:14<5863 2015 99 253 https://doi.org/10.1007/s00253-014-6328-9 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00253-014-6328-9 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Unzueta Ugutz Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut NATIONALLICENCE 700 1- Vázquez Felicitas Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut NATIONALLICENCE 700 1- Accardi Giulia Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut NATIONALLICENCE 700 1- Mendoza Rosa Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut NATIONALLICENCE 700 1- Toledo-Rubio Verónica Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut NATIONALLICENCE 700 1- Giuliani Maria Dipartimento di Scienze Biochimiche, Facoltà di Medicina e Chirurgia, Università degli studi di Palermo, 90127, Palermo, Italy aut NATIONALLICENCE 700 1- Sannino Filomena Dipartimento di Scienze Biochimiche, Facoltà di Medicina e Chirurgia, Università degli studi di Palermo, 90127, Palermo, Italy aut NATIONALLICENCE 700 1- Parrilli Ermenegilda Dipartimento di Scienze Biochimiche, Facoltà di Medicina e Chirurgia, Università degli studi di Palermo, 90127, Palermo, Italy aut NATIONALLICENCE 700 1- Abasolo Ibane CIBER de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Barcelona, Spain aut NATIONALLICENCE 700 1- Schwartz Jr Simo CIBER de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Barcelona, Spain aut NATIONALLICENCE 700 1- Tutino Maria Dipartimento di Scienze Biochimiche, Facoltà di Medicina e Chirurgia, Università degli studi di Palermo, 90127, Palermo, Italy aut NATIONALLICENCE 700 1- Villaverde Antonio Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut NATIONALLICENCE 700 1- Corchero José Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut NATIONALLICENCE 700 1- Ferrer-Miralles Neus Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain aut NATIONALLICENCE 773 0- Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/14(2015-07-01), 5863-5874 0175-7598 99:14<5863 2015 99 253