Shielding of protein-boronate interactions during boronate chromatography of neoglycoproteins

A method for separating glycoproteins on a boronate column under conditions which suppress the interactions between the protein moiety and the boronic acid ligand has been developed. A model system consisting of non-glycosyla ted chymotrypsin and maltose-modified chymotrypsin (cht-mal) was utilised i n the investigations. Chymotrypsin was chosen as the model protein because of its known interaction with boronate. By coupling maltose to chymotrypsin , a neoglycoprotein was created which has the property of binding to the af finity matrix both via the; protein moiety and via the carbohydrate residue s. The introduction of a so-called shielding reagent into the buffer soluti ons during chromatography resulted in the prevention of the protein-boronat e interactions while the carbohydrate-boronate interaction was little influ enced. Different types of, mainly low-molecular-mass, polyhydroxyl chemical s were screened in order to correlate the shielding efficiency to the chemi cal structure of the investigated compounds. Polyhydroxyl chemicals with a conformation that allows the formation of tridentate complexes with the bor onate anion provided the highest shielding efficiencies. (C) 2001 Elsevier Science B.V. All rights reserved.