title: Investigation of chromatography and polymer/salt aqueous two-phase processes for downstream processing development of recombinant phenylalanine dehydrogenase.
abstract: This work presents a comprehensive study between the polymer/salt aqueous two-phase systems (ATPS) and chromatography process for downstream processing of recombinant Bacillus badius phenylalanine dehydrogenase (PheDH). First, the partitioning behavior of recombinant PheDH in polyethylene glycol (PEG)/K2HPO4 ATPS was examined. For comparative purpose, a classical chromatographic protocol was performed as well. Investigation of chromatography and ATPS procedures revealed that the ATPS comprising of 9% (w/w) PEG-6000, 16% (w/w) K2HPO4 and 16% (w/w) KCl with pH of 8.0, volume ratio (V ( R )) of 0.25, temperature of 25 degrees C and 40% (w/w) cell lysate ensured the most favorable approach for PheDH downstream process. A specific activity of 4,231.4 U/mg, a yield of 96.7% and a recovery of 162.0% were obtained. Furthermore, the shorter process time (4 vs. 48 h) and the lower total cost (4 vs. 20 euro) were additionally features that confirmed the suitability of proposed technique.
authors.affiliation: Department of Biochemistry, Pasteur Institute of Iran, 13164, Tehran, Iran.
authors.forename: Eskandar
authors.initials: E
authors.lastname: Omidinia
authors.forename: Hamid
authors.initials: H
authors.lastname: Shahbaz Mohamadi
authors.forename: Rassoul
authors.initials: R
authors.lastname: Dinarvand
authors.forename: Heshmat-Allah
authors.initials: HA
authors.lastname: Taherkhani
mesh_terms: D000594:Amino Acid Oxidoreductases / Q000737:chemistry*; D001709:Biotechnology / Q000379:methods*; D002845:Chromatography / Q000379:methods*; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D011092:Polyethylene Glycols / Q000737:chemistry; D011108:Polymers / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry*; D012492:Salts / Q000737:chemistry; D014867:Water / Q000737:chemistry
chemical_list: D011108:Polymers; D011994:Recombinant Proteins; D012492:Salts; D014867:Water; D011092:Polyethylene Glycols; D000594:Amino Acid Oxidoreductases; C022342:phenylalanine oxidase