Purification of glucose-6-phosphate dehydrogenase from baker's yeast in aqueous two-phase systems with free triazine dyes as affinity ligands.
To improve the selectivity of glucose-6-phosphate dehydrogenase (G6PDH) extraction by an aqueous two-phase system, a simple and inexpensive affinity aqueous two-phase system using unbound reactive triazine dyes as ligands was introduced. In a polyethylene glycol (PEG)/hydroxypropyl starch (PES) system, the unbound free triazine dyes, Cibacron Blue F3GA and Procion Red HE3B, partitioned unevenly in the top PEG-rich phase and thus showed an affinity effect on G6PDH, but no influence on hexokinase. The various parameters investigated were pH of the system, buffers, molecular weight of PEG, and ligand type and concentration. A two-step affinity extraction process was established for the purification of G6PDH from baker's yeast. The total yield of G6PDH was 66.9% and purification factor was 2.35.
Xu
Yan
Y
Department of Chemical Engineering, Hebei Institute of Technology, Tangshan, Hebei, 063009, P.R. China.
Vitolo
Michele
M
de Albuquerque
Cristina Northfleet
CN
Pessoa
Adalberto
A
D002846:Chromatography, Affinity; D004396:Coloring Agents; D005954:Glucosephosphate Dehydrogenase / Q000302:isolation & purification* / Q000378:metabolism*; D006593:Hexokinase / Q000378:metabolism; D007700:Kinetics; D008024:Ligands; D008970:Molecular Weight; D011092:Polyethylene Glycols; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D013056:Spectrophotometry, Ultraviolet; D014227:Triazines
D004396:Coloring Agents; D008024:Ligands; D014227:Triazines; D011092:Polyethylene Glycols; D005954:Glucosephosphate Dehydrogenase; D006593:Hexokinase