[Fusion expression of PLA2 gene from Lapemis hardwickii in E. coli].
The gene encoding PLA2(PLA2-9) from Lapemis hardwickii Gray venom was cloned to the 3' and of the thioredoxin gene (HP-trxA and trxA) in plasmid pthioHisC and pTRX to construct the pThioHisC-PLA2, and pTRX-PLA2 fusion expression vector. The fusion protein of PLA2 can be expressed in the two different systems induced by IPTG at 25 degrees C, but the expression level and the solubility of the fusion protein in pTRX were better than that in pThioHisC. The expressed product in the two systems were purified by immobilized metal-chelate affinity chromatography. The Trx-PLA2 fusion protein with over 85% purity was obtained and HP-Trx-PLA2 fusion protein can not be purified since it dose not exhibit affinity to the medium. So, the pTRX-PLA2 vector system was established for the large-scale expression and purification.
Yang
Wenli
W
National Opening Laboratory for Marine Functional Genomics Research, School of Life Science, Zhongshan University, Guangzhou 510275, China.
Ye
Lanting
L
Zou
Lan
L
Wang
Feng
F
Xu
Anlong
A
D000818:Animals; D004546:Elapid Venoms / Q000737:chemistry*; D017815:Elapidae; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D005822:Genetic Vectors; D010741:Phospholipases A / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D054467:Phospholipases A2; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D013879:Thioredoxins / Q000235:genetics*
D004546:Elapid Venoms; D011993:Recombinant Fusion Proteins; D013879:Thioredoxins; D010741:Phospholipases A; D054467:Phospholipases A2