Expression, purification, and characterization of recombinant lumbrokinase PI239 in Escherichia coli.
Lumbrokinase (LK) is an important fibrinolytic enzyme derived from earthworms. It has been found that LK is composed of a group of isoenzymes. To construct and express the mature peptide of LK PI239 in Escherichia coli, we amplified and optimized the gene of LK which was then cloned into the prokaryotic expression vector pET-22b(-). The recombinant LK (rLK) protein was expressed as inclusion bodies and we have developed a purification process of rLK from these inclusion bodies. A step-down urea concentration strategy was applied to the rLK renaturation process. The purified and renatured rLK apparently ameliorated the conditions of the model thrombosis rats used, and may be developed into a therapeutic agent for thrombotic-associated diseases.
Xu
Zhe-rong
ZR
Department of Geriatrics, First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou 310003, China.
Yang
Yun-mei
YM
Gui
Qi-feng
QF
Zhang
Li-na
LN
Hu
Lin
L
D000818:Animals; D010450:Endopeptidases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism* / Q000627:therapeutic use; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D002479:Inclusion Bodies / Q000201:enzymology; D007527:Isoenzymes / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008297:Male; D009835:Oligochaeta / Q000201:enzymology*; D011897:Random Allocation; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism* / Q000627:therapeutic use; D013927:Thrombosis / Q000139:chemically induced / Q000188:drug therapy; D010959:Tissue Plasminogen Activator / Q000009:adverse effects
D007527:Isoenzymes; D011994:Recombinant Proteins; D010450:Endopeptidases; D010959:Tissue Plasminogen Activator; C083421:lumbrokinase