title: Microcalorimetric studies of the effects on the interactions of human recombinant interferon-alpha2a.
abstract: The applicability of the physical stability of protein solution monitored by isothermal titration calorimetry (ITC) was evaluated. The second virial coefficient, b2, derived from the dilution enthalpies of protein solution measured by ITC under various experimental conditions was studied. The protein applied in this work is human recombinant interferon-alpha2a (hrIFN-alpha2a), which is a commercial drug applied for the treatment of virus-infected diseases. The results obtained were used to predict the possibility of hrIFN-alpha2a aggregation, and the prediction can be further confirmed by size-exclusion chromatography (SEC). Various factors affecting the stability of protein solution were investigated, for example, temperature, salts, surfactants, and mechanical stress. Specifically, the results show that the dilution enthalpy of hrIFN-alpha2a increased with increasing temperature and NaCl concentration, while b2 decreased, indicating that the attraction between hrIFN-alpha2a molecules was enhanced under these conditions. On studying the effect of mechanical stress, the data obtained reveals that the introduction of centrifugal or vortex force strengthened the attractive forces between hrIFN-alpha2a molecules. These implications were supported by SEC data, demonstrating that the amount of aggregated hrIFN-alpha2a was increased. As a consequence, the methodologies presented in this investigation offer a possibility of monitoring the physical stability of protein solution at various stages of recovery, purification as well as the development of appropriate drug storage formulations.
authors.affiliation: Department of Life Sciences, National Central University, Jhongli City, Taoyuan County 320, Taiwan. slhuang@cc.ncu.edu.tw
authors.forename: Shir-Ly
authors.initials: SL
authors.lastname: Huang
authors.forename: Fu-Yung
authors.initials: FY
authors.lastname: Lin
authors.forename: Chih-Ping
authors.initials: CP
authors.lastname: Yang
mesh_terms: D002151:Calorimetry; D002850:Chromatography, Gel; D006863:Hydrogen-Ion Concentration; D000077190:Interferon alpha-2; D016898:Interferon-alpha / Q000737:chemistry*; D009566:Nitrates / Q000494:pharmacology; D011994:Recombinant Proteins; D012965:Sodium Chloride / Q000494:pharmacology; D013314:Stress, Mechanical; D013501:Surface-Active Agents / Q000494:pharmacology; D013696:Temperature; D013816:Thermodynamics
chemical_list: D000077190:Interferon alpha-2; D016898:Interferon-alpha; D009566:Nitrates; D011994:Recombinant Proteins; D013501:Surface-Active Agents; D012965:Sodium Chloride; C031618:sodium nitrate