[Renaturation, purification and antigenicity identification of recombinant protein of Cysticercus cellulosae expressed in Escherichia coli].
To obtain the recombinant 18 kD protein with high purity and normal bioactivity of Cysticercus cellulosae (rCE18), E. coli cells with the rCE18 were disrupted ultra-sonically, and the inclusion bodies were washed with a solution containing 0.2% deoxycholic acid sodium (DOC)and 2% DOC, respectively. Then they were denatured with 0.9% sodium lauroyl sarcosine (SKL) followed by dialysis and gel filtration to refold and purify the target protein. At the same time, this method was compared with GST-FF affinity chromatography and recovering from SDS-PAGE gel. Biological activity of purified rCE18 was analyzed with indirect ELISA, and the purity of the products was identified using SDS-PAGE. The purity of refolded inclusion bodies exceeded 60% and the total recovery of activated protein rCE18 was about 41.3%. The specificity of rCE18 reached up to 97.2% using indirect ELISA. An effective way for purifying and refolding rCE18 expressed in E. coli as inclusion bodies was established, rCE18 with higher purity and activity was obtained, which has the potential for developing diagnosis methods of porcine cysticercosis.
Zhang
Shaohua
S
Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy ofAgricultural Sciences, Lanzhou 730046, China.
Jia
Wanzhong
W
Luo
Xuenong
X
Jing
Zhizhong
Z
Wu
Guohua
G
Zheng
Yadong
Y
Guo
Aijiang
A
Cai
Xuepeng
X
D000818:Animals; D000947:Antigens, Helminth / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D002850:Chromatography, Gel; D003552:Cysticercus / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D002479:Inclusion Bodies / Q000378:metabolism; D020673:Protein Renaturation; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*
D000947:Antigens, Helminth; D011994:Recombinant Proteins