Neutral salts exhibit very marked differences as eluants of proteins from affinity columns. We observe: (i) that the relative potencies of neutral salts as eluants are independent of the protein or the affinity ligand in the systems studied, (ii) that the absolute salt concentration necessary to elute any given protein bound to the affinity matrix is proportional to the algebraic sum of a set of elution coefficients defined herein for the separate ions present in the solution, and (iii) that the proportionality between elution potency and elution coefficient is a function of the affinity of the protein for the immobilized ligand. Given the concentration ofone neutral salt required for elution of a protein ofinterest from an affinity column, the elution capability of any neutral salt at any temperature can be quantitatively predicted for that protein. Accordingly, application and elution protocols for affinity chromatography can be designed to optimize the yield.and fold purification of proteins.While affinity chromatography has expedited the purification ofa wide range ofproteins, use ofa substrate or substrate analog as a selective eluant is often very expensive, particularly in the large-scale purification procedures required for acquisition of useful quantities ofproteins oflow natural abundance. For such situations various neutral salt solutions have been utilized as an eluant (1, 2), often resulting in a fold purification inferior to that with a specific ligand as the eluant. We find that various properties of the solutions under study correlate closely with the concentration of salt sufficient to remove enzyme bound to an affinity system, with the most useful correlation being found with the B coefficient in the equation postulated by Jones and Dole (3). This correlation can be exploited to optimize the fold purification and yield achieved by affinity chromatography by neutral salt elution.
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