title: Protein Internal Dynamics Associated With Pre-System Glass Transition Temperature Endothermic Events: Investigation of Insulin and Human Growth Hormone by Solid State Hydrogen/Deuterium Exchange.
abstract: Lyophilized proteins are generally stored below their glass transition temperature (Tg) to maintain long-term stability. Some proteins in the (pure) solid state showed a distinct endotherm at a temperature well below the glass transition, designated as a pre-Tgendotherm. The pre-Tgendothermic event has been linked with a transition in protein internal mobility. The aim of this study was to investigate the internal dynamics of 2 proteins, insulin and human growth hormone (hGH), both of which exhibit the pre-Tgendothermic event with onsets at 50°C-60°C. Solid state hydrogen/deuterium (H/D) exchange of both proteins was characterized by Fourier transform infrared spectroscopy over a temperature range from 30°C to 80°C. A distinct sigmoidal transition in the extent of H/D exchange had a midpoint of 56.1 ± 1.2°C for insulin and 61.7 ± 0.9°C for hGH, suggesting a transition to greater mobility in the protein molecules at these temperatures. The data support the hypothesis that the pre-Tgevent is related to a transition in internal protein mobility associated with the protein dynamical temperature. Exceeding the protein dynamical temperature is expected to activate protein internal motion and therefore may have stability consequences.
authors.affiliation: Department of Pharmaceutical Sciences, University of Connecticut, Storrs, Connecticut 06269.
authors.forename: Rui
authors.initials: R
authors.lastname: Fang
authors.affiliation: Department of Pharmaceutical Sciences, University of Connecticut, Storrs, Connecticut 06269.
authors.forename: Pawel J
authors.initials: PJ
authors.lastname: Grobelny
authors.affiliation: Department of Pharmaceutical Sciences, University of Connecticut, Storrs, Connecticut 06269; Institute of Materials Science, University of Connecticut, Storrs, Connecticut 06269.
authors.forename: Robin H
authors.initials: RH
authors.lastname: Bogner
authors.affiliation: Department of Pharmaceutical Sciences, University of Connecticut, Storrs, Connecticut 06269; Institute of Materials Science, University of Connecticut, Storrs, Connecticut 06269. Electronic address: michael.pikal@uconn.edu.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Pikal
mesh_terms: D041961:Deuterium Exchange Measurement / Q000379:methods*; D005898:Glass / Q000737:chemistry*; D019382:Human Growth Hormone / Q000737:chemistry*; D006801:Humans; D007328:Insulin / Q000737:chemistry*; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods; D044366:Transition Temperature
chemical_list: D007328:Insulin; D019382:Human Growth Hormone
keywords: FTIR; amorphous; calorimetry (DSC); freeze drying/lyophilization; glass transition; mobility; protein structure; proteins; solid state stability