Metal chelate affinity precipitation:: purification of (HiS)6-tagged lactate dehydrogenase using poly(vinylimidazole-co-N-isopropylacrylamide) copolymers

Affinity precipitation is a highly selective bioseparation method, which has been successfully used in the purification of various enzymes/proteins. The separation of (His)(6)-tagged lactate dehydrogenase ((His)(6) LDH) was carried through affinity precipitation technique using thermoresponsive copolymer poly(vinylimidazole-co-N-isopropyl-acrylamide) (poly(VI-NIPAM)). Nearly quantitative recovery of (His)(6)-LDH has been achieved using Cu(II)- and Ni(II)-loaded poly(VI-NIPAM) with Cu(II)-poly(VI-NIPAM) being more efficient in enzyme precipitation than Ni(H)-poly(VI-NIPAM). Optimal precipitation of (His)(6)-LDH was achieved at pH 6 and pH 6.5-7.0 with Cu(II)-poly(VI-NIPAM) and Ni(II)-poly(VI-NIPAM), respectively. Furthermore, the enzyme was purified by affinity precipitation upto 3.5- and 3.1-fold with recoveries of 95 and 82% using Cu(II)- and Ni(II)-loaded copolymers, respectively. (C) 2003 Elsevier Science Inc. All rights reserved.