title: Isolation and characterization of a monomethioninesulfoxide variant of interferon alpha-2b.
abstract: OBJECTIVE
  To isolate and characterize a monomethioninesulfoxide variant of the commercially available therapeutic protein interferon alpha-2b.
  
  
  METHODS
  The methionine (Met)-oxidized variant was isolated by reverse-phase high performance liquid chromatography and characterized by SDS-PAGE, peptide mapping and mass spectrometric analysis of the trypsin/V8-generated peptide fragments. The biological and immunological activities of the isolated variant were also evaluated.
  
  
  RESULTS
  The rHuIFN alpha-2b variant was found to contain a Met sulfoxide residue at position 111 of the rHuIFN alpha-2b molecule. The far-UV CD spectra showed a slight loss of alpha-helical content and an increase in the beta-sheet contribution. The CD spectra indicate that both chromatographic conditions and Met oxidation contribute to the observed secondary structure changes. Both interferon alpha-2b main component and its methionine-oxidized variant showed different reactivity to monoclonal antibodies employed in immunoassays for the protein.
  
  
  CONCLUSIONS
  A monomethioninesulfoxide rHuIFN alpha-2b variant was found to be present in the rHuIFN alpha-2b bulk drug substance in solution. The Met(111) residue was identified as Met sulfoxide by comparative tryptic/V8 mapping and mass spectrometric analysis. Nevertheless, the oxidation of the Met(111) residue did not seem to have a detectable effect on the biological activity of the molecule.
authors.affiliation: Schering-Plough Research Institute, Kenilworth, New Jersey 07033, USA.
authors.forename: G
authors.initials: G
authors.lastname: Gitlin
authors.forename: A
authors.initials: A
authors.lastname: Tsarbopoulos
authors.forename: S T
authors.initials: ST
authors.lastname: Patel
authors.forename: W
authors.initials: W
authors.lastname: Sydor
authors.forename: B N
authors.initials: BN
authors.lastname: Pramanik
authors.forename: S
authors.initials: S
authors.lastname: Jacobs
authors.forename: L
authors.initials: L
authors.lastname: Westreich
authors.forename: S
authors.initials: S
authors.lastname: Mittelman
authors.forename: J N
authors.initials: JN
authors.lastname: Bausch
mesh_terms: D000595:Amino Acid Sequence; D000998:Antiviral Agents / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D000077190:Interferon alpha-2; D016898:Interferon-alpha / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D013058:Mass Spectrometry; D008715:Methionine / Q000031:analogs & derivatives* / Q000737:chemistry; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D011994:Recombinant Proteins; D013056:Spectrophotometry, Ultraviolet
chemical_list: D000998:Antiviral Agents; D000077190:Interferon alpha-2; D016898:Interferon-alpha; D011994:Recombinant Proteins; D008715:Methionine; C013111:methionine sulfoxide