Binding of higher plant NADH-dependent nitrate reductase to different triazine dyes

Affinity partitioning in an aqueous two-phase system consisting of dextran, polyethylene glycol and triazine dye-substituted polyethylene glycol was applied to investigate the affinity of NADH-dependent nitrate reductases isolated from tobacco, barley and cucumber to six triazine dyes. The alteration of the partition coefficient of nitrate reductases in the presence and absence of dye-polyethylene glycol conjugates provided qualitative data for the affinity of the enzymes to the triazine dyes. Cibacron Blue F3G-A, widely used as a ligand for purification of nitrate reductase by affinity chromatography had a low affinity for higher plant nitrate reductases compared to other triazine dyes. The strength of interaction between dye and enzyme was not dependent on the enzyme source but on the dye used. The influence of NADH on the binding of triazine dyes to nitrate reductase was also investigated.