1. The effect of urea and trimethyl-N-amine oxide (TMAO) upon the heteroassociation of alpha-chymotrypsin (chy) and soybean trypsin inhibitor (STI) was studied by measurement of fluorescence depolarization. STI was labeled with Alexa-488 and titrated with chy. The dependence of the rotational relaxation time of labeled STI upon the concentration of added chy was interpreted in the context of a model in which chy could bind to two independent sites on STI with approximately equal affinity. Titrations were conducted in the presence of various added concentrations of urea and TMAO, and it was found that while added urea tended to inhibit the association of chy and STI, TMAO enhanced the association. Analysis of the dependence of results upon the concentrations of added urea and TMAO showed that the two effects, although in opposite directions, were additive. (D. Wu) 2. The overall effect of a concentrated crowding agent upon a model for self-association of a dilute reactant was analyzed as the sum of the effects of both excluded volume and attractive interactions between the crowding agent and the reactant species. (A. Minton)

Project Start
Project End
Budget Start
Budget End
Support Year
6
Fiscal Year
2013
Total Cost
$202,850
Indirect Cost
City
State
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Hoppe, Travis; Minton, Allen P (2015) An equilibrium model for the combined effect of macromolecular crowding and surface adsorption on the formation of linear protein fibrils. Biophys J 108:957-66
Beg, Ilyas; Minton, Allen P; Hassan, Imtaiyaz et al. (2015) Thermal Stabilization of Proteins by Mono- and Oligosaccharides: Measurement and Analysis in the Context of an Excluded Volume Model. Biochemistry 54:3594-603
Minton, Allen P (2013) Quantitative assessment of the relative contributions of steric repulsion and chemical interactions to macromolecular crowding. Biopolymers 99:239-44
Wu, Di; Minton, Allen P (2013) Compensating effects of urea and trimethylamine-N-oxide on the heteroassociation of α-chymotrypsin and soybean trypsin inhibitor. J Phys Chem B 117:3554-9
Fernández, Cristina; Minton, Allen P (2011) Effect of nonadditive repulsive intermolecular interactions on the light scattering of concentrated protein-osmolyte mixtures. J Phys Chem B 115:1289-93
Nagarajan, Sureshbabu; Amir, Dan; Grupi, Asaf et al. (2011) Modulation of functionally significant conformational equilibria in adenylate kinase by high concentrations of trimethylamine oxide attributed to volume exclusion. Biophys J 100:2991-9
Jiao, Ming; Li, Hong-Tao; Chen, Jie et al. (2010) Attractive protein-polymer interactions markedly alter the effect of macromolecular crowding on protein association equilibria. Biophys J 99:914-23
Fodeke, Adedayo A; Minton, Allen P (2010) Quantitative characterization of polymer-polymer, protein-protein, and polymer-protein interaction via tracer sedimentation equilibrium. J Phys Chem B 114:10876-80
Tsao, Douglas; Minton, Allen P; Dokholyan, Nikolay V (2010) A didactic model of macromolecular crowding effects on protein folding. PLoS One 5:e11936
Zhou, Huan-Xiang; Rivas, German; Minton, Allen P (2008) Macromolecular crowding and confinement: biochemical, biophysical, and potential physiological consequences. Annu Rev Biophys 37:375-97