TY - JOUR
T1 - Metal chelate affinity precipitation: purification of (His)6-tagged lactate dehydrogenase using poly(vinylimidazole-co-N-isopropylacrylamide)copolymers
AU - Kumar, Ashok
AU - Khalil, Ashraf
AU - Galaev, Igor
AU - Mattiasson, Bo
PY - 2003
Y1 - 2003
N2 - 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(II)-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.
AB - 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(II)-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.
KW - Affinity precipitation
KW - Lactate dehydrogenase
KW - Thermoresponsive copolymers
KW - Metal affinity
U2 - 10.1016/S0141-0229(03)00092-9
DO - 10.1016/S0141-0229(03)00092-9
M3 - Article
SN - 0141-0229
VL - 33
SP - 113
EP - 117
JO - Enzyme and Microbial Technology
JF - Enzyme and Microbial Technology
IS - 1
ER -