Metal Ion-promoted Dye-ligand Chromatography

Abstract

Over the last decade, triazine dyes have received widespread attention as tools in protein purification technology (Lowe et al., 1981). It is only in more recent years, however, that the potential of multivalent metal ions in promoting protein binding to triazine dye affinity adsorbents was recognised (Hughes et al., 1982a). Previously, little reference had been made to the role of metal ions in triazine dye affinity chromatography. Only the influence of Mg2+ had received some attention, for example, as a requirement to achieve maximum binding of yeast hexokinase to immobilised Procion Green H4-G (Clonis et al., 1981). Enzyme was subsequently eluted using buffers from which the metal ions had been omitted. High concentrations of Mg2+ were also used during the purification of isoleucyl and glycyl tRNA synthetases on Blue Dextran—Sepharose (Moe and Piszkiewicz, 1976). In this case the isoleucyl enzyme was eluted with an ATP gradient after Mg2+ had been removed by washing with the equilibration buffer, while glycyl tRNA synthetase was eluted using a decreasing Mg2+ gradient. The binding of frog liver carbamyl phosphate synthetase I (Mori and Cohen, 1978) and isocitrate dehydrogenase (Nagaoka et al., 1977) to immobilised Cibacron Blue F3G-A had also been shown to be influenced by Mg2+; these enzymes were eluted with Mg2+-free buffer and the chelating agent EDTA, respectively.