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1. Zhongguo Xue Xi Chong Bing Fang Zhi Za Zhi. 2011 Dec;23(6):653-8.

[Expression and characterization of recombinant Sj23HD-HSA fusion protein in
Saccharomyces cerevisiae].

[Article in Chinese]

Zhang W, Yu CX, Yang JL, Feng W, Yin XR, Song LJ, Wang J, Qian CY, Ke XD.

Department of Novel Biotechnique, Jiangsu Institute of Parasitic Diseases, Key
Laboratory on Technology for Parasitic Diseases Prevention and Control, Ministry
of Health, Wuxi 214064, China.

OBJECTIVES: To prepare the fusion protein of large hydrophilic domain of 23 kDa
membrane protein of Schistosoma japonicum and the mature peptide of human serum
albumin (Sj23HD-HSA) and investigate its immunoreactivity.
METHODS: A fusion protein gene encoding Sj23HD-HSA fusion protein was prepared by
overlapping PCR, which was confirmed by TA cloning and DNA sequencing. The fusion
gene of Sj23HD-HSA was directionally subcloned into yeast expression plasmid
pWX530 to construct a recombinant plasmid Sj23HD-HSA/pWX530. The transformants of
Saccharomyces cerevisiae containing the recombinant plasmid Sj23HD-HSA/pWX530
were screened on leu deficient SD medium after yeast competent cells were
transformed with recombinant plasmid. The excretive Sj23HD-HSA protein was
expressed by culturing the yeast transformants at 30 degrees C for 1 week, and
the protein component of culture supernatant was analyzed by SDS-PAGE. Sj23HD-HSA
fusion protein was purified through Ion Exchange Chromatography. The
immunoreactivity of recombinant Sj23HD-HSA fusion protein was determined by
Western blotting with sera of schistosomiasis, clonorchiasis and healthy.
RESULTS: The gene encoding the Sj23HD-HSA fusion protein was constructed
successfully, which was confirmed by DNA sequencing. The yeast transformants
containing plasmid Sj23HD-HSA/pWX530 could express the excretive Sj23HD-HSA
fusion protein without inducing. The results of Western blotting indicated
Sj23HD-HSA could be recognized by the sera of schistosomiasis, but could not be
recognized by the sera of clonorchiasis and healthy respectively.
CONCLUSIONS: Sj23HD-HSA fusion protein with good immune reactivity is prepared
successfully, which will be a potential antigen for schistosomiasis
immunodiagnosis.

PMID: 22379820 [PubMed - in process]