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Dye‐ligand membranes as selective adsorbents for rapid purification of enzymes: A case study
Author(s) -
Champluvier B.,
Kula M.R.
Publication year - 1992
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.260400106
Subject(s) - chemistry , chromatography , membrane , elution , adsorption , yield (engineering) , microporous material , affinity chromatography , ligand (biochemistry) , enzyme , ethylene glycol , filtration (mathematics) , yeast , biochemistry , organic chemistry , materials science , receptor , statistics , mathematics , metallurgy
Abstract A new adsorbent for the selective binding of enzymes, in the form of microporous membranes carrying triazine dyes as pseudo‐affinity ligand, has been implemented in the recovery of glucose‐6‐phosphate dehydrogenase from yeast. A detailed investigation of the process parameters has been performed. In the adsorption step, the contact time for binding G6PDH could be reduced down to 0.25 s without significant decrease of the capture efficiency. Hence, fast filtration allowed to isolate G6PDH from a dilute extract (1.6 μg G6PDH · mL −1 ), where the enzyme accounted for 1% of the proteins. The yield of the selective elution step using NADP was only 70% at best. It could be improved to near 100% by supplementing the eluent with ethylene glycol, without loss of selectivity. A Scale‐up of the cross‐section of the membrane by a factor of 40 allowed to purify 1140 U from 0.6 L extract from 1% to 57% purity with 82% yield, within 10 minutes. The case study presented here demonstrates the applicability of general‐purpose membrane adsorbents for the purification of enzymes.