title: Recombinant production of a single-chain antibody fragment in Pseudoalteromonas haloplanktis TAC125.
abstract: Recombinant protein production in cold-adapted bacteria has proved to be a valuable option to overcome solubility concerns often came up in conventional expression hosts. ScFvs are examples of "difficult proteins" due to their tendency to form inclusion bodies when expressed in Escherichia coli. In this paper, the recombinant production of a single-chain antibody (ScFvOx) in the psychrophilic bacterium Pseudoalteromonas haloplanktis TAC125 is reported. The expression vector for the ScFvOx production was designed to address the recombinant protein in the periplasmic space and to allow the formation of the antibody disulphide bonds. For periplasmic export, two different export mechanisms were evaluated. By combining the genetic tools available for recombinant protein expression in psychrophilic hosts with an ad hoc medium and fermentation modality and optimised expression conditions at low temperatures, we obtained the highest yield of soluble and epitope-binding ScFvOx reported so far by conventional prokaryotic expression. The observed proficiency of the Antarctic bacterium to produce recombinant antibody fragments was related to the unusually high number of genes encoding peptidyl prolyl cis-trans isomerases found in P. haloplanktis TAC125 genome, making this bacterium the host of choice for the recombinant production of this protein class.
authors.affiliation: Dipartimento di Scienze Chimiche, Università di Napoli "Federico II", Via Cinthia, 4, 80126, Naples, Italy, maria.giuliani@ymail.com.
authors.forename: Maria
authors.initials: M
authors.lastname: Giuliani
authors.forename: Ermenegilda
authors.initials: E
authors.lastname: Parrilli
authors.forename: Filomena
authors.initials: F
authors.lastname: Sannino
authors.forename: Gennaro Antonio
authors.initials: GA
authors.lastname: Apuzzo
authors.forename: Gennaro
authors.initials: G
authors.lastname: Marino
authors.forename: Maria Luisa
authors.initials: ML
authors.lastname: Tutino
mesh_terms: D003080:Cold Temperature; D005822:Genetic Vectors; D021381:Protein Transport; D031161:Pseudoalteromonas / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D057127:Single-Chain Antibodies / Q000235:genetics / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D057127:Single-Chain Antibodies