title: Effect of arginine on pre-nucleus stage of interferon beta-1b aggregation.
abstract: Understanding the mechanism of aggregation of a therapeutic protein would not only ease the manufacturing processing but could also lead to a more stable finished product. Aggregation of recombinant interferon (IFNβ-1b) was studied by heating, oxidizing, or seeding of unformulated monomeric solution. The formation of aggregates was monitored by dynamic light scattering (DLS) and UV spectroscopy. The autocatalytic monomer loss model was used to fit the data on aggregation rates. The influence of pre-nucleation on aggregation step was demonstrated by inducing the liquid samples containing a monomer form of folded IFNβ-1b by heat and also an oxidizing agent. Results tend to suggest that the nucleus includes a single protein molecule which has been probably deformed. Seeding tests showed that aggregation of IFNβ-1b was probably initiated when 1.0% (w/w) of monomers converted to nucleus form. Chemiluminescence spectroscopy analysis of the sample indicated the generation of 3.0 μM of hydrogen peroxide (H2O2) during nucleation stage of IFNβ-1b aggregation. Arginine with a concentration of 200 mM was sufficient to suppress aggregation of IFNβ-1b by decreasing the rate of pre-nucleation step. We proposed the formation of pre-nucleus structures prior to nucleation as the mechanism of aggregation of IFNβ-1b. Furthermore, we have showed the positive anti-aggregation effect of arginine on pre-nucleation step.
authors.affiliation: Biotechnology Group, Department of Chemical Engineering, Tarbiat Modares University, Tehran, Iran.
authors.forename: Ahmad
authors.initials: A
authors.lastname: Fazeli
authors.forename: Mohadeseh
authors.initials: M
authors.lastname: Haji-Abdolvahab
authors.forename: Seyed Abbas
authors.initials: SA
authors.lastname: Shojaosadati
authors.forename: Huub
authors.initials: H
authors.lastname: Schellekens
authors.forename: Khosro
authors.initials: K
authors.lastname: Khalifeh
authors.forename: Ali Akbar
authors.initials: AA
authors.lastname: Moosavi-Movahedi
authors.forename: Mohammad Reza
authors.initials: MR
authors.lastname: Fazeli
mesh_terms: D000998:Antiviral Agents / Q000737:chemistry* / Q000494:pharmacology; D001120:Arginine / Q000737:chemistry*; D045744:Cell Line, Tumor; D003588:Cytopathogenic Effect, Viral / Q000187:drug effects; D004686:Encephalomyocarditis virus / Q000187:drug effects; D005079:Excipients / Q000737:chemistry*; D006801:Humans; D006861:Hydrogen Peroxide / Q000737:chemistry; D000068576:Interferon beta-1b; D016899:Interferon-beta / Q000737:chemistry* / Q000494:pharmacology; D007700:Kinetics; D008027:Light; D008956:Models, Chemical; D010084:Oxidation-Reduction; D066329:Protein Aggregates; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry; D012542:Scattering, Radiation; D013056:Spectrophotometry, Ultraviolet; D013678:Technology, Pharmaceutical / Q000379:methods
chemical_list: D000998:Antiviral Agents; D005079:Excipients; D066329:Protein Aggregates; D011994:Recombinant Proteins; D000068576:Interferon beta-1b; D016899:Interferon-beta; D001120:Arginine; D006861:Hydrogen Peroxide