An improved method for purification of recombinant truncated heme oxygenase-1 by expanded bed adsorption and gel filtration.
Recombinant truncated human heme oxygenase-1 (hHO-1) expressed in Escherichia coli was efficiently separated and purified from feedstock by DEAE-ion exchange expanded bed adsorption. Protocol optimization of hHO-1 on DEAE adsorbent resulted in adsorption in 0 M NaCl and elution in 150 mM NaCl at a pH of 8.5. The active enzyme fractions separated from the expanded bed column were further purified by a Superdex 75 gel filtration step. The specific hHO-1 activity increased from 0.82 +/- 0.05 to 24.8 +/- 1.8 U/mg during the whole purification steps. The recovery and purification factor of truncated hHO-1 of the whole purification were 72.7 +/- 4.7 and 30.2 +/- 2.3%, respectively. This purification process can decrease the demand on the preparation of feedstock and simplify the purification process.
Hu
Hong-Bo
HB
Key Laboratory of Microbial Metabolism, Ministry of Education, College of Life Science and Biotechnology, Shanghai Jiao Tong University, Shanghai, People's Republic of China. hbhu@sjtu.edu.cn
Wang
Wei
W
Han
Ling
L
Zhou
Wen-Pu
WP
Zhang
Xue-Hong
XH
D002852:Chromatography, Ion Exchange / Q000379:methods*; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D051547:Heme Oxygenase-1 / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D006801:Humans; D011994:Recombinant Proteins / Q000302:isolation & purification* / Q000378:metabolism; D014462:Ultrafiltration / Q000379:methods*
D011994:Recombinant Proteins; D051547:Heme Oxygenase-1