title: Antibiotic free selection for the high level biosynthesis of a silk-elastin-like protein.
abstract: Silk-elastin-like proteins (SELPs) are a family of genetically engineered recombinant protein polymers exhibiting mechanical and biological properties suited for a wide range of applications in the biomedicine and materials fields. They are being explored as the next generation of biomaterials but low productivities and use of antibiotics during production undermine their economic viability and safety. We have developed an industrially relevant, scalable, fed-batch process for the high level production of a novel SELP in E. coli in which the commonly used antibiotic selection marker of the expression vector is exchanged for a post segregational suicide system, the separate-component-stabilisation system (SCS). SCS significantly augments SELP productivity but also enhances the product safety profile and reduces process costs by eliminating the use of antibiotics. Plasmid content increased following induction but no significant differences in plasmid levels were discerned when using SCS or the antibiotic selection markers under the controlled fed-batch conditions employed. It is suggested that the absence of competing plasmid-free cells improves host cell viability and enables increased productivity with SCS. With the process developed, 12.8 g L-1purified SELP was obtained, this is the highest SELP productivity reported to date and clearly demonstrates the commercial viability of these promising polymers.
authors.affiliation: Centre of Molecular and Environmental Biology (CBMA), Department of Biology, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal.
authors.forename: Mário
authors.initials: M
authors.lastname: Barroca
authors.affiliation: Centre of Molecular and Environmental Biology (CBMA), Department of Biology, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal.
authors.forename: Paulo
authors.initials: P
authors.lastname: Rodrigues
authors.affiliation: Biosystems and Integrative Sciences Institute (BioISI), Plant Functional Biology Centre, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal.
authors.forename: Rómulo
authors.initials: R
authors.lastname: Sobral
authors.affiliation: Biosystems and Integrative Sciences Institute (BioISI), Plant Functional Biology Centre, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal.
authors.forename: M Manuela R
authors.initials: MM
authors.lastname: Costa
authors.affiliation: Centre of Molecular and Environmental Biology (CBMA), Department of Biology, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal.
authors.forename: Susana R
authors.initials: SR
authors.lastname: Chaves
authors.affiliation: Centre of Molecular and Environmental Biology (CBMA), Department of Biology, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal.
authors.forename: Raul
authors.initials: R
authors.lastname: Machado
authors.affiliation: Centre of Molecular and Environmental Biology (CBMA), Department of Biology, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal.
authors.forename: Margarida
authors.initials: M
authors.lastname: Casal
authors.affiliation: Centre of Molecular and Environmental Biology (CBMA), Department of Biology, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal.
authors.forename: Tony
authors.initials: T
authors.lastname: Collins
mesh_terms: D001709:Biotechnology / Q000379:methods*; D004549:Elastin / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000254:growth & development / Q000378:metabolism*; D005827:Genetics, Microbial / Q000379:methods*; D042822:Genomic Instability; D050296:Microbial Viability; D010957:Plasmids; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D012641:Selection, Genetic
chemical_list: D011994:Recombinant Proteins; D004549:Elastin