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1 May 2005 Association of Eppin with Semenogelin on Human Spermatozoa
Zengjun Wang, E. E. Widgren, P. Sivashanmugam, M. G. O'Rand, R. T. Richardson
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Eppin (SPINLW1; GeneID, 57119) is a single-copy gene encoding a cysteine-rich protein found only in the testis and epididymis, which contains both Kunitz-type and WAP-type four disulfide core protease inhibitor consensus sequences. This study demonstrates that, in seminal plasma and on human spermatozoa following ejaculation, Eppin is bound to semenogelin I (Sg). Six different experimental approaches: 1) immunoprecipitation from spermatozoa and seminal plasma with anti-Eppin, 2) colocalization in semen and spermatozoa, 3) incubation of recombinant Eppin (rEppin) and rSg and immunoprecipitation with either anti-Eppin or anti-Sg, 4) far-Western blotting of Eppin and Sg, 5) Saturation binding of 125I-Sg to Eppin, which is competed by unlabeled Sg, and 6) direct binding of 125I-Sg to Eppin on a blot, all demonstrate that Eppin and Sg bind to each other. To study the specificity of binding, recombinant fragments of Eppin and Sg were made and demonstrate that the Eppin75–133 C-terminal fragment binds the Sg164–283 fragment containing the only cysteine in human Sg I (Cys-239). Reduction and carboxymethylation of Cys239 blocks binding of 125I-rEppin, indicating that a disulfide bond may be necessary for Eppin binding. The physiological significance of the Eppin-semenogelin complex bound on the surface of ejaculate spermatozoa lies in its ability to provide antimicrobial activity for spermatozoa, which has been reported for both Eppin and semenogelin-derived peptides, and in its ability to provide for the survival and preparation of spermatozoa for fertility in the female reproductive tract.

Zengjun Wang, E. E. Widgren, P. Sivashanmugam, M. G. O'Rand, and R. T. Richardson "Association of Eppin with Semenogelin on Human Spermatozoa," Biology of Reproduction 72(5), 1064-1070, (1 May 2005).
Received: 21 September 2004; Accepted: 1 December 2004; Published: 1 May 2005

male reproductive tract
seminal vesicles
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