Rapid measurement of protein osmotic second virial coefficients by self-interaction chromatography.
Weak protein interactions are often characterized in terms of the osmotic second virial coefficient (B(22)), which has been shown to correlate with protein phase behavior, such as crystallization. Traditional methods for measuring B(22), such as static light scattering, are too expensive in terms of both time and protein to allow extensive exploration of the effects of solution conditions on B(22). In this work we have measured protein interactions using self-interaction chromatography, in which protein is immobilized on chromatographic particles and the retention of the same protein is measured in isocratic elution. The relative retention of the protein reflects the average protein interactions, which we have related to the second virial coefficient via statistical mechanics. We obtain quantitative agreement between virial coefficients measured by self-interaction chromatography and traditional characterization methods for both lysozyme and chymotrypsinogen over a wide range of pH and ionic strengths, yet self-interaction chromatography requires at least an order of magnitude less time and protein than other methods. The method thus holds significant promise for the characterization of protein interactions requiring only commonly available laboratory equipment, little specialized expertise, and relatively small investments of both time and protein.
Tessier
Peter M
PM
Center for Molecular and Engineering Thermodynamics, Department of Chemical Engineering, University of Delaware, Newark, Delaware 19716, USA.
Lenhoff
Abraham M
AM
Sandler
Stanley I
SI
D000818:Animals; D055592:Biophysical Phenomena; D001703:Biophysics; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D002645:Chickens; D002845:Chromatography / Q000379:methods*; D002919:Chymotrypsinogen / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D008027:Light; D015233:Models, Statistical; D009113:Muramidase / Q000737:chemistry; D009995:Osmosis; D011485:Protein Binding; D012542:Scattering, Radiation
D002919:Chymotrypsinogen; D009113:Muramidase