title: Effects of molecular crowding by saccharides on alpha-chymotrypsin dimerization.
abstract: Given the importance of protein complexes as therapeutic targets, it is necessary to understand the physical chemistry of these interactions under the crowded conditions that exist in cells. We have used sedimentation equilibrium to quantify the enhancement of the reversible homodimerization of alpha-chymotrypsin by high concentrations of the osmolytes glucose, sucrose, and raffinose. In an attempt to rationalize the osmolyte-mediated stabilization of the alpha-chymotrypsin homodimer, we have used models based on binding interactions (transfer-free energy analysis) and steric interactions (excluded volume theory) to predict the stabilization. Although transfer-free energy analysis predicts reasonably well the relatively small stabilization observed for complex formation between cytochrome c and cytochrome c peroxidase, as well as that between bobtail quail lysozyme and a monoclonal Fab fragment, it underestimates the sugar-mediated stabilization of the alpha-chymotrypsin dimer. Although predictions based on excluded volume theory overestimate the stabilization, it would seem that a major determinant in the observed stabilization of the alpha-chymotrypsin homodimer is the thermodynamic nonideality arising from molecular crowding by the three small sugars.
authors.affiliation: Department of Chemistry, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
authors.forename: Chetan N
authors.initials: CN
authors.lastname: Patel
authors.forename: Schroeder M
authors.initials: SM
authors.lastname: Noble
authors.forename: Gresham T
authors.initials: GT
authors.lastname: Weatherly
authors.forename: Ashutosh
authors.initials: A
authors.lastname: Tripathy
authors.forename: Donald J
authors.initials: DJ
authors.lastname: Winzor
authors.forename: Gary J
authors.initials: GJ
authors.lastname: Pielak
mesh_terms: D002918:Chymotrypsin / Q000737:chemistry*; D019281:Dimerization; D004735:Energy Transfer; D005947:Glucose / Q000737:chemistry*; D007700:Kinetics; D011485:Protein Binding; D011887:Raffinose / Q000737:chemistry*; D013395:Sucrose / Q000737:chemistry*
chemical_list: D013395:Sucrose; D002918:Chymotrypsin; C117608:alpha-chymotrypsin; D005947:Glucose; D011887:Raffinose