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Cytoplasmic expression, antibody production, and characterization of the novel zinc finger protein 637

  • Applied genetics and molecular biotechnology
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Abstract

Zinc finger protein 637 (zfp637), belonging to the Kruppel-like protein family, comprises one atypical C2H2 and six consecutive typical zinc finger motifs. Based on the structural characterization of zfp637 and its location in the cell nucleus, we predict that zfp637 might function as a DNA-binding protein to regulate gene transcription. However, the absence of both a purified zfp637 protein and any commercial antibody for detecting it in cells and tissues has limited functional studies of zfp637 to date. Here, we developed and optimized an expression system by fusing zfp637 with glutathione S-transferase (GST) to achieve a maximal yield of soluble GST-zfp637 fusion protein in Escherichia coli BL21(DE3) cells. The yield was about 10 mg/l of the original bacterial culture. The recombinant GST-zfp637 fusion protein was purified and used for polyclonal antibody production in rabbits. In addition, we developed a method to remove the anti-GST antibody component and obtained a highly purified anti-zfp637 antibody, as demonstrated by an enzyme-linked immunosorbent assay. Western blotting showed that the anti-zfp637 antibody recognized not only the recombinant zfp637 protein but also endogenous zfp637 in several cell lines. The protein was localized mainly in the cell nucleus by immunofluorescence and immunohistochemistry. The expression levels of zfp637 mRNA and protein were significantly increased in NIH3T3 cells treated with 200 μM of H2O2 in a time-dependent manner. The recombinant GST-zfp637 fusion protein and our purified anti-zfp637 antibody will help in elucidating the function of zfp637.

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Acknowledgments

This work was supported by items (31070675) from the National Natural Science Foundation of China. It was also supported by Sichuan Province Foundation (2010JY0055) from the Science and Technology Department of Sichuan Province.

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The authors declare that they have no conflict of interest.

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Authors and Affiliations

  1. Division of Geriatrics, State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, 1# Keyuan 4 Road, Gaopeng Avenue, Hi-tech Development, Chengdu, 610041, Sichuan, People’s Republic of China

    Kai Li, Yuyan Wei, Jie Zhang, Jun Li, Bo Gao & Ping Lin

  2. Department of Pediatric Surgery, West China Hospital, Sichuan University, 37 Guo Xue Xiang, Chengdu, Sichuan, 610041, People’s Republic of China

    Lugang Huang

  3. Division of Cancer Biotherapy, State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan, People’s Republic of China

    Yuquan Wei

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  1. Kai Li
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  2. Yuyan Wei
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  3. Jie Zhang
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  4. Jun Li
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Correspondence to Ping Lin.

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Li, K., Wei, Y., Zhang, J. et al. Cytoplasmic expression, antibody production, and characterization of the novel zinc finger protein 637. Appl Microbiol Biotechnol 97, 741–749 (2013). https://doi.org/10.1007/s00253-012-4235-5

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  • DOI: https://doi.org/10.1007/s00253-012-4235-5

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