title: Strain and process development for the production of human cytokines in Hansenula polymorpha.
abstract: The early status of strain development for the production of interleukin (IL)-6, IL-8, IL-10, and interferon (IFN) gamma is described. The general approach to generating such strains was to amplify gene sequences encoding the mature forms of the various cytokines by PCR from commercially available cDNA sources. The design of the amplificates allowed an in-frame fusion to an MFalpha1 leader segment contained in two basic expression vectors, pFPMT121-MFalpha1 and pTPSMT-MFalpha1. The two vectors differ in that one harbors the methanol-inducible FMD promoter and the other the constitutive TPS1 promoter as control elements for heterologous gene expression. The most advanced process development example is that of IFNalpha-2a. Here, the MOX promoter derived from another key gene of methanol metabolism is used for expression control. The successful development of a production process for Hansenula polymorpha-derived IFNalpha-2a is summarized. This was achieved by combining genetic engineering of suitable production strains with improved processing capabilities for the secreted cytokine, and by purification procedures from cultures grown in yeast extract-peptone-glycerol-based media.
authors.affiliation: Rhein Biotech GmbH, Eichsfelder Strasse 11, 40595 Düsseldorf, Germany.
authors.forename: Adelheid
authors.initials: A
authors.lastname: Degelmann
authors.forename: Frank
authors.initials: F
authors.lastname: Müller
authors.forename: Heike
authors.initials: H
authors.lastname: Sieber
authors.forename: Volker
authors.initials: V
authors.lastname: Jenzelewski
authors.forename: Manfred
authors.initials: M
authors.lastname: Suckow
authors.forename: Alexander W M
authors.initials: AW
authors.lastname: Strasser
authors.forename: Gerd
authors.initials: G
authors.lastname: Gellissen
mesh_terms: D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D016207:Cytokines / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D005285:Fermentation; D005822:Genetic Vectors; D006801:Humans; D008969:Molecular Sequence Data; D010843:Pichia / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
chemical_list: D016207:Cytokines; D011994:Recombinant Proteins