Dye—ligand affinity chromatography on continuous beds

✍ Scribed by Jamil Mohammad; Asadullah Zeerak; Stellan Hjertén

• Publisher: John Wiley and Sons
• Year: 1995
• Tongue: English
• Weight: 408 KB
• Volume: 9
• Category: Article
• ISSN: 0269-3879
• DOI: 10.1002/bmc.1130090205

✦ Synopsis

Continuous beds derivatized with three triazine dyes (Cibacron Blue 3G-A, Pricon Red HE3B and Pricon Red H-3B) were used for chromatographic purification of dehydrogenases from yeast enzyme concentrate. All three columns, which were prepared by a very simple and cost-effective method, provided strong binding of glucose-6phosphate dehydrogenase and lactate dehydrogenase. The Cibacron Blue 3G-A column showed high affinity for alcohol dehydrogenase. Under the same conditions, the Pricon Red HE3B column showed a lower affinity and the Pricon Red H-3B column showed none. The adsorbed dehydrogenases were eluted specifically from the columns in high yields (71-113% by desorption with the coenzymes NADP, NADH and N A D respectively).

Non-specific binding of human serum albumin and transferrin to these columns was also investigated. Enzyme assays and analyses by capillary electrophoresis showed that the continuous beds derivatized with triazine dyes gave a high degree of purification.