[Expression and biology identification of the human epididymis-specific gene ESC42 in E. coli].
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.


CONCLUSIONS
Obtaining high purity of rhESC42 protein may lay a foundation for the study of its functions.
Shen
Xiao-Fang
XF
Central Laboratory of Yantai Yuhuangding Hospital, Shandong Stem Cell Engineering Research Center, Yantai, Shandong 264000, China. sxfcym@yahoo.com.en
Li
Jian-Yuan
JY
Wang
Hai-Yan
HY
Xue
Jiang-Nan
JN
Cao
Qi-Zhi
QZ
Lian
Pei-Wen
PW
Chen
Hui
H
Xie
Mei-Na
MN
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D003001:Cloning, Molecular; D023082:Defensins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D004822:Epididymis / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D015723:Gene Library; D006801:Humans; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000276:immunology
D000911:Antibodies, Monoclonal; C437905:DEFB118 protein, human; D023082:Defensins; D011993:Recombinant Fusion Proteins