Protein self-association induced by macromolecular crowding: a quantitative analysis by magnetic relaxation dispersion.
In the presence of high concentrations of inert macromolecules, the self-association of proteins is strongly enhanced through an entropic, excluded-volume effect variously called macromolecular crowding or depletion attraction. Despite the predicted large magnitude of this universal effect and its far-reaching biological implications, few experimental studies of macromolecular crowding have been reported. Here, we introduce a powerful new technique, fast field-cycling magnetic relaxation dispersion, for investigating crowding effects on protein self-association equilibria. By recording the solvent proton spin relaxation rate over a wide range of magnetic field strengths, we determine the populations of coexisting monomers and decamers of bovine pancreatic trypsin inhibitor in the presence of dextran up to a macromolecular volume fraction of 27%. Already at a dextran volume fraction of 14%, we find a 30-fold increase of the decamer population and 510(5)-fold increase of the association constant. The analysis of these results, in terms of a statistical-mechanical model that incorporates polymer flexibility as well as the excluded volume of the protein, shows that the dramatic enhancement of bovine pancreatic trypsin inhibitor self-association can be quantitatively rationalized in terms of hard repulsive interactions.
Snoussi
Karim
K
Department of Biophysical Chemistry, Lund University, SE-22100 Lund, Sweden. karim.snoussi@port.ac.uk
Halle
Bertil
B
D000818:Animals; D007611:Aprotinin / Q000737:chemistry; D001703:Biophysics / Q000379:methods*; D002417:Cattle; D003911:Dextrans / Q000737:chemistry; D019277:Entropy; D006859:Hydrogen / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D046911:Macromolecular Substances / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008280:Magnetics; D008958:Models, Molecular; D015233:Models, Statistical; D011108:Polymers / Q000737:chemistry; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry*; D011522:Protons; D012997:Solvents; D013696:Temperature; D013816:Thermodynamics
D003911:Dextrans; D046911:Macromolecular Substances; D011108:Polymers; D011506:Proteins; D011522:Protons; D012997:Solvents; D006859:Hydrogen; D007611:Aprotinin