Authors : Z. Sharifi , B. yakhchali and M.S. Shahrabadi
Abstract: Rotavirus nonstructural glycoprotein, NSP4 has been proposed as the first viral enterotoxin capable of inducing diarrhea and a target for vaccine development. In order to study biological role of NSP4, a cDNA from human rotavirus (Wa strain) RNA segment 10 was amplified by PCR, using two specific and cloned into cloning vector pBS-KS(+) and subsequently into pQE-30 expression plasmid. Expression of NSP4 was demonstrated by SDS-PAGE,Western blot and ELISA using polyclonal antibody against NSP4 from SA11 infected BSC1 cells. The recombinant protein was purified by an affinity chromatography on Nickle NTA-Agarose column (Qiagen). A polyclonal antiserum against purified recombinant NSP4 was raised in Rabbit; which was reacted with NSP4 in BSC1 cells infected with SA11 rotavirus. Intraperitoneally inoculation of NSP4 caused diarrhea in BALB/c suckling mice indicating its biological activity. Intraperitoneal and oral inoculation of NSP4 antiserum significantly decreased diarrhea disease. These results indicated successful expression and purification of the full-length biologically active, NSP4 of human rotavirus Wa strain in E. coli and showed that antibody against it was able to protect against simian rotavirus diarrhea in neonatal mice.
Z. Sharifi , B. yakhchali and M.S. Shahrabadi , 2005. Expression and One Step Purification of The Full-length Biologically Active, Nsp4 Of Human Rotavirus Wa Strain. International Journal of Molecular Medicine and Advance Sciences, 1: 206-212.