title: Use of high-gradient magnetic fishing for reducing proteolysis during fermentation.
abstract: Proteolysis during fermentation may have a severe impact on the yield and quality of a secreted product. In the current study, we demonstrate the use of high-gradient magnetic fishing (HGMF) as an efficient alternative to the more conventional methods of preventing proteolytic degradation. Bacitracin-linked magnetic affinity adsorbents were employed directly in a fermenter during Bacillus licheniformis cultivation to remove trace amounts of unwanted proteases. The constructed magnetic adsorbents had excellent, highly specific binding characteristics in the fermentation broth (K(d)  = 1.94 micromolar; Q(max)  = 222.8 mg/g), which obeyed the Langmuir isotherm and had rapid binding kinetics (equilibrium in <300 s). When applied directly in shake-flask cultures or in a 1-L fermenter and then removed by HGMF, the degradation of the model protein bovine serum albumin was stopped. The adsorbents could be recycled and reused during the same fermentation to remove freshly produced proteases, extending the life of the model protein in the fermenter. HGMF may provide an efficient method of stabilizing heterologous proteins produced in cultivation processes.
authors.affiliation: Center for Microbial Biotechnology, Technical University of Denmark, Lyngby, Denmark.
authors.forename: Trine L
authors.initials: TL
authors.lastname: Maury
authors.forename: Kim E
authors.initials: KE
authors.lastname: Ottow
authors.forename: Jesper
authors.initials: J
authors.lastname: Brask
authors.forename: John
authors.initials: J
authors.lastname: Villadsen
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Hobley
mesh_terms: D000327:Adsorption; D000818:Animals; D001407:Bacillus / Q000737:chemistry / Q000378:metabolism; D018533:Biomass; D001709:Biotechnology / Q000295:instrumentation / Q000379:methods*; D002417:Cattle; D005285:Fermentation; D007700:Kinetics; D059346:Magnets; D010447:Peptide Hydrolases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D011485:Protein Binding; D059748:Proteolysis; D012710:Serum Albumin, Bovine / Q000737:chemistry / Q000378:metabolism
chemical_list: D012710:Serum Albumin, Bovine; D010447:Peptide Hydrolases