EXPRESSION AND CHARACTERIZATION OF AN EPIDIDYMIS-SPECIFIC GENE

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Abstract
A truncated cDNA coding a rabbit epididymal protein (BE-20) was identified in a previous study. In the present study the full-length cDNA was isolated by the method of rapid amplification of cDNA ends. The BE-20 cDNA consisted of 585 bp with a poly(A) tail of 26 residues and an open reading frame composed of 369 bp encoding a deduced polypeptide containing 123 amino acid residues with a calculated molecular mass of 13 kDa. The N-terminus-contained a leucine-rich segment. BE-20 cDNA has about 76.8% homology with the HE4 gene of human epididymis. Northern blot analysis of mRNAs prepared from 17 different human tissues was performed using as probe a 0.5-kb DNA fragment corresponding to a segment of BE-20 cDNA. Positive reaction was elicited only with epididymal mRNA. A DNA fragment corresponding to a section of the open reading frame of BE-20 cDNA was cloned in Escherichia coli under the control of the T7 promoter. The cellular content of the expressed recombinant protein comprised about 55% of the total protein. The chromatographically purified bacterial product migrated as a single band with an estimated M(r) of 15 kDa on analysis by SDS-PAGE. In conclusion, BE-20 cDNA is expressed only in the epididymis. It is structurally related to the four-disulfide core family of extracellular proteinase inhibitors and may be involved in sperm maturation.