title: Optimization of the expression of recombinant human activin A in the yeast Pichia pastoris.
abstract: We report a new procedure to express recombinant human activin A using the methanolic yeast, Pichia pastoris. Optimization of culture procedures has involved comprehensive examination of the effects of culture vessel shape, volume of broth in the induction and expression cultures, methanol concentration, culturing temperature, and pH of the expression cultures. After this optimization, as well as modification of the native cleavage sites, a laboratory scale procedure has been established which routinely produced 2-10 mg/L amounts of this vital growth factor in the highly efficient, eukaryotic yeast system. This system avoids the need to produce this protein and similar TGF-beta proteins in mammalian cell lines which, in addition to being costly, produce many native binding partners of these cystine knot proteins, a factor which can dramatically affect yields of the target protein.
authors.affiliation: ARC Special Research Centre for Green Chemistry, Monash University, Clayton, VIC 3800, Australia.
authors.forename: Dale
authors.initials: D
authors.lastname: Fredericks
authors.forename: Robert
authors.initials: R
authors.lastname: Clay
authors.forename: Tracy
authors.initials: T
authors.lastname: Warner
authors.forename: Anne
authors.initials: A
authors.lastname: O'Connor
authors.forename: David M
authors.initials: DM
authors.lastname: de Kretser
authors.forename: Milton T W
authors.initials: MT
authors.lastname: Hearn
mesh_terms: D028341:Activins / Q000096:biosynthesis* / Q000235:genetics; D001483:Base Sequence; D018533:Biomass; D018929:Cell Culture Techniques / Q000379:methods*; D003001:Cloning, Molecular / Q000379:methods; D006801:Humans; D006863:Hydrogen-Ion Concentration; D000432:Methanol / Q000737:chemistry; D008969:Molecular Sequence Data; D010843:Pichia / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D013696:Temperature
chemical_list: D011994:Recombinant Proteins; C422405:activin A; D028341:Activins; D000432:Methanol