Food Biochemistry and Food Processing

8 Enzyme Engineering and Technology 201

products, cofactors, inhibitors, or allosteric effectors
are all potential candidates as long as they have
higher affinity for the macromolecule than for the
immobilized ligand.
Dye-ligand affinity chromatographyrepresents
a powerful affinity-based technique for enzyme and
protein purification (Clonis et al. 2000, Labrou
2002, Labrou et al. 2004). The technique has gained
broad popularity due to its simplicity and wide
applicability to purifying a variety of proteins. The
employed dyes used as affinity ligands are commer-
cial textile chlorotriazine polysulfonated aromatic
molecules, which are usually termed triazine dyes
(Fig. 8.15). Such dye ligands have found wide appli-
cation in the research market over the past 20 years
as low specificity general affinity ligands to purify

enzymes such as oxidoreductases, decarboxylases,

glycolytic enzymes, nucleases, hydrolases, lyases,

synthetases, and transferases (Scopes 1987). Anthr-

aquinone triazine dyes are probably the most widely

used dye ligands in enzyme and protein purification.

The triazine dye Cibacron Blue 3GA (Fig. 8.15),

especially, has been widely exploited as an affinity

chromatographic tool to separate and purify a vari-

ety of proteins (Scopes 1987). With the aim of in-

creasing the specificity of dye ligands, the biomimet-

ic dye-ligand concept was introduced. According to

this concept, new dyes that mimic natural ligands of

the targeted proteins are designed by substituting the

terminal 2-aminobenzene sulfonate moiety of the

dye Cibacron Blue 3GA (CB3GA) with a substrate-

mimetic moiety (Clonis et al. 2000; Labrou 2002,

Figure 8.15.Structure of several representative triazine dyes: (A)Cibacron Blue 3GA, (B) Procion Red HE-3B,
(C) Procion Rubine MX-B.