Zhonghua nan ke xue = National journal of andrology · 2005
[Expression and biology identification of the human epididymis-specific gene ESC42 in E. coli].
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Shen Xiao-Fang, Li Jian-Yuan, Wang Hai-Yan, Xue Jiang-Nan, Cao Qi-Zhi, Lian Pei-Wen, Chen Hui, Xie Mei-Na
Аннотация
OBJECTIVE: To provide materials for the study of the function of ESC42 protein specifically expressed in the human epididymis.
METHODS: The ESC42 gene was amplified from the human epididymis cDNA library by PCR and then cloned into prokaryotic expression vector pGEX-4T-1, expressed and purified by recombinant DNA techniques. The specificity of ESC42 protein was identified by Western blot and MALDI-TOF-MS. The database was searched by Ms-Fit.
RESULTS: The recombinant plasmid expressed a Mr 38 x 10(3) fusion protein in E. coli at a level of 30% of the total protein, and the purity was as high as 99%. The ESC42 protein was identified by ESC42 monoclonal antibody and its molecular weight was 11 978.12, tested by MALDI-TOF-MS. The peptide mass fingerprint analysis showed that the coverage rate of the sequence reached 48% with 100% matching. The motif scan in Prosite database reveal that ESC42 belonged to the beta-defensin family and had antibacterial activity.
CONCLUSION: Obtaining high purity of rhESC42 protein may lay a foundation for the study of its functions.
METHODS: The ESC42 gene was amplified from the human epididymis cDNA library by PCR and then cloned into prokaryotic expression vector pGEX-4T-1, expressed and purified by recombinant DNA techniques. The specificity of ESC42 protein was identified by Western blot and MALDI-TOF-MS. The database was searched by Ms-Fit.
RESULTS: The recombinant plasmid expressed a Mr 38 x 10(3) fusion protein in E. coli at a level of 30% of the total protein, and the purity was as high as 99%. The ESC42 protein was identified by ESC42 monoclonal antibody and its molecular weight was 11 978.12, tested by MALDI-TOF-MS. The peptide mass fingerprint analysis showed that the coverage rate of the sequence reached 48% with 100% matching. The motif scan in Prosite database reveal that ESC42 belonged to the beta-defensin family and had antibacterial activity.
CONCLUSION: Obtaining high purity of rhESC42 protein may lay a foundation for the study of its functions.