title: Modeling of protein-anion exchange resin interaction for the human growth hormone charge variants.
abstract: Modeling ion exchange chromatography (IEC) behavior has generated significant interest because of the wide use of IEC as an analytical technique as well as a preparative protein purification process; indeed there is a need for better understanding of what drives the unique behavior of protein charge variants. We hypothesize that a complex protein molecule, which contains both hydrophobic and charged moieties, would interact strongly with an in silico designed resin through charged electrostatic patches on the surface of the protein. In the present work, variants of recombinant human growth hormone that mimic naturally-occurring deamidation products were produced and characterized in silico. The study included these four variants: rhGH, N149D, N152D, and N149D/N152D. Poisson-Boltzmann calculations were used to determine surface electrostatic potential. Metropolis Monte Carlo simulations were carried out with the resulting variants to simulate IEC systems, examining the free energy of the interaction of the protein with an in silico anion exchange column represented by polylysine polypeptide. The results show that the charge variants have different average binding energies and the free energy of interaction can be used to predict the retention time for the different variants.
authors.affiliation: Department of Late Stage Pharmaceutical Development, Genentech Inc. (member of Roche), South San Francisco, CA 94080, USA. Electronic address: lapelosm@gene.com.
authors.forename: Mauro
authors.initials: M
authors.lastname: Lapelosa
authors.affiliation: Department of Early Stage Pharmaceutical Development, Genentech Inc. (member of Roche), South San Francisco, CA 94080, USA.
authors.forename: Thomas W
authors.initials: TW
authors.lastname: Patapoff
authors.affiliation: Department of Late Stage Pharmaceutical Development, Genentech Inc. (member of Roche), South San Francisco, CA 94080, USA.
authors.forename: Isidro E
authors.initials: IE
authors.lastname: Zarraga
mesh_terms: D000837:Anion Exchange Resins / Q000737:chemistry*; D019382:Human Growth Hormone / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D056004:Molecular Dynamics Simulation; D009010:Monte Carlo Method; D016297:Mutagenesis, Site-Directed; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D055672:Static Electricity
chemical_list: D000837:Anion Exchange Resins; D011994:Recombinant Proteins; D019382:Human Growth Hormone
keywords: IEC; Molecular simulations; Monte Carlo; rhGH–column interaction