title: Denatured state aggregation parameters derived from concentration dependence of protein stability.
abstract: Protein aggregation is a major issue affecting the long-term stability of protein preparations. Proteins exist in equilibrium between the native and denatured or partially denatured conformations. Often denatured or partially denatured conformations are prone to aggregate because they expose to solvent the hydrophobic core of the protein. The aggregation of denatured protein gradually shifts the protein equilibrium toward increasing amounts of denatured and ultimately aggregated protein. Recognizing and quantitating the presence of denatured protein and its aggregation at the earliest possible time will bring enormous benefits to the identification and selection of optimal solvent conditions or the engineering of proteins with the best stability/aggregation profile. In this article, a new approach that allows simultaneous determination of structural stability and the amount of denatured and aggregated protein is presented. This approach is based on the analysis of the concentration dependence of the Gibbs energy (ΔG) of protein stability. It is shown that three important quantities can be evaluated simultaneously: (i) the population of denatured protein, (ii) the population of aggregated protein, and (iii) the fraction of denatured protein that is aggregated.
authors.affiliation: Department of Biology, Johns Hopkins University, Baltimore, MD 21218, USA.
authors.forename: Arne
authors.initials: A
authors.lastname: Schön
authors.affiliation: Department of Biology, Johns Hopkins University, Baltimore, MD 21218, USA.
authors.forename: Benjamin R
authors.initials: BR
authors.lastname: Clarkson
authors.affiliation: Department of Biology, Johns Hopkins University, Baltimore, MD 21218, USA.
authors.forename: Rogelio
authors.initials: R
authors.lastname: Siles
authors.affiliation: Unchained Labs, Norton, MA 02766, USA.
authors.forename: Patrick
authors.initials: P
authors.lastname: Ross
authors.affiliation: Unchained Labs, Norton, MA 02766, USA.
authors.forename: Richard K
authors.initials: RK
authors.lastname: Brown
authors.affiliation: Department of Biology, Johns Hopkins University, Baltimore, MD 21218, USA; Unchained Labs, Norton, MA 02766, USA. Electronic address: ef@jhu.edu.
authors.forename: Ernesto
authors.initials: E
authors.lastname: Freire
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry*; D001120:Arginine / Q000737:chemistry; D024402:Carbonic Anhydrase II / Q000737:chemistry*; D002417:Cattle; D000068818:Cetuximab / Q000737:chemistry*; D004355:Drug Stability; D004795:Enzyme Stability; D006358:Hot Temperature / Q000009:adverse effects; D007202:Indicators and Reagents / Q000737:chemistry; D008958:Models, Molecular; D009994:Osmolar Concentration; D010974:Platelet Aggregation; D066329:Protein Aggregates; D011487:Protein Conformation; D011489:Protein Denaturation; D055550:Protein Stability; D012995:Solubility; D013816:Thermodynamics; D000068878:Trastuzumab / Q000737:chemistry*; D014508:Urea / Q000737:chemistry
chemical_list: D000970:Antineoplastic Agents; D007202:Indicators and Reagents; D066329:Protein Aggregates; D014508:Urea; D001120:Arginine; D024402:Carbonic Anhydrase II; D000068878:Trastuzumab; D000068818:Cetuximab
keywords: Denatured state aggregation; Isothermal chemical denaturation; Protein conformational equilibrium; Thermodynamic linkage equilibrium and aggregation