title: Cultivation of Pichia pastoris carrying the scFv anti LDL (-) antibody fragment. Effect of preculture carbon source.
abstract: Antibodies and antibody fragments are nowadays among the most important biotechnological products, and Pichia pastoris is one of the most important vectors to produce them as well as other recombinant proteins. The conditions to effectively cultivate a P. pastoris strain previously genetically modified to produce the single-chain variable fragment anti low density lipoprotein (-) under the control of the alcohol oxidase promoter have been investigated in this study. In particular, it was evaluated if, and eventually how, the carbon source (glucose or glycerol) used in the preculture preceding cryopreservation in 20% glycerol influences both cell and antibody fragment productions either in flasks or in bioreactor. Although in flasks the volumetric productivity of the antibody fragment secreted by cells precultured, cryopreserved and reactivated in glycerol was 42.9% higher compared with cells precultured in glucose, the use of glycerol in bioreactor led to a remarkable shortening of the lag phase, thereby increasing it by no less than thrice compared to flasks. These results are quite promising in comparison with those reported in the literature for possible future industrial applications of this cultivation, taking into account that the overall process time was reduced by around 8h.
authors.affiliation: Universidade de São Paulo, Departamento Bioquímico - Tecnologia Farmacêutica, São Paulo, SP, Brazil.
authors.forename: Cesar Andres Diaz
authors.initials: CAD
authors.lastname: Arias
authors.affiliation: Universidade Rural Federal de Pernambuco, Departamento de Morfologia e Fisiologia Animal, Recife, PE, Brazil.
authors.forename: Daniela de Araujo Viana
authors.initials: DAV
authors.lastname: Marques
authors.affiliation: Universidade de São Paulo, Departamento Bioquímico - Tecnologia Farmacêutica, São Paulo, SP, Brazil.
authors.forename: Luciana Pellegrini
authors.initials: LP
authors.lastname: Malpiedi
authors.affiliation: Universidade Nacional de Brasíilia, Departamento de Biologia Celular, Laboratório de Imunologia Molecular, Brasília, DF, Brazil.
authors.forename: Andrea Queiroz
authors.initials: AQ
authors.lastname: Maranhão
authors.affiliation: Universidade de São Paulo, Departamento de Análises Clínicas e Toxicologia, São Paulo, SP, Brazil.
authors.forename: Dulcineia Abdalla Saes
authors.initials: DAS
authors.lastname: Parra
authors.affiliation: University of Genoa, Department of Civil, Chemical and Environmental Engineering, Genoa, Italy. Electronic address: converti@unige.it.
authors.forename: Attilio
authors.initials: A
authors.lastname: Converti
authors.affiliation: Universidade de São Paulo, Departamento Bioquímico - Tecnologia Farmacêutica, São Paulo, SP, Brazil.
authors.forename: Adalberto Pessoa
authors.initials: AP
authors.lastname: Junior
mesh_terms: D000906:Antibodies / Q000235:genetics / Q000378:metabolism*; D002244:Carbon / Q000378:metabolism*; D003470:Culture Media / Q000737:chemistry / Q000378:metabolism; D005285:Fermentation; D005990:Glycerol / Q000378:metabolism; D007218:Industrial Microbiology / Q000379:methods*; D008077:Lipoproteins, LDL / Q000276:immunology; D010843:Pichia / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D057127:Single-Chain Antibodies / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D000906:Antibodies; D003470:Culture Media; D008077:Lipoproteins, LDL; D011994:Recombinant Proteins; D057127:Single-Chain Antibodies; D002244:Carbon; D005990:Glycerol
keywords: Carbon source; Cryopreservation; Expression; Pichia pastoris; scFv antibody fragment