Epididymal protease inhibitor (EPPIN) is an androgen-regulated sperm-binding protein (Yenugu S et al. 2004). EPPIN is expressed specifically in the testis and epididymis (Richardson RT et al. 2001). EPPIN coats the surface of human testicular and epididymal spermatozoa as part of a protein complex containing lactotransferrin (LTF) and clusterin (CLU) (Wang Z et al. 2007; Paasch U et al. 2011). During ejaculation, semenogelin 1 (SEMG1) binds to EPPIN in the complex (Wang Z et al. 2005; 2007; O'Rand MG et al, 2009; 2011; Silva EJ et al. 2012). The components of the complex, EPPIN, LTF and SEMG1-derived peptides have been shown to possess microbicidal activity. EPPIN exhibited dose- and time-dependent antibacterial activity against E. coli (Yenugu S et al. 2004). While the activity of EPPIN was relatively insensitive to salt, it was completely lost on reduction and alkylation of Cys residues of EPPIN, indicating the importance of disulfide bonds (Yenugu S et al. 2004). Furthermore, EPPIN was able to induce morphological alterations of E. coli membranes as shown by scanning electron microscopy and measuring bacterial respiratory electron transport (Yenugu S et al. 2004; McCrudden MT et al. 2008). LTF is a multifunctional protein that is best known for its ability to bind iron, which eventually led to the discovery of its antibacterial activity. In addition, LTF has demonstrated potent antiviral, antifungal and antiparasitic activity towards a broad spectrum of species (Legrand D et al. 2008). SEMG1-derived peptides were found in the cationic fraction of seminal plasma and showed high levels of antibacterial activity against E. coli and B. megaterium (Bourgeon F et al. 2004; Edstrom AM et al. 2008). The EPPIN protein complex provides antimicrobial activity on the sperm surface. In addition, the binding of SEMG1 to EPPIN during ejaculation inhibits sperm progressive motility, possibly by disturbing the regulation of intracellular pH, resulting in the loss of calcium (O'Rand MG et al, 2009; O'Rand MG & Widgren EE 2012). In parallel, EPPIN inhibits the digestion of SEMG1 by the serine protease prostate specific antigen (PSA), which results in the modulation of semen liquefaction, further prolonging the inhibitory effects of SEMG1 on sperm motility (Wang Z et al. 2005; Wang ZJ et al. 2008; Silva EJ et al. 2013).

EPPIN has two potential protease inhibitory domains: a whey acid protein (WAP)-type four disulfide core (WFDC) domain and a Kunitz domain (Richardson RT et al. 2001; McCrudden MT et al. 2008). While both domains of EPPIN are involved in the protein's antibacterial activity, only the Kunitz domain is required for selective protease inhibition (McCrudden MT et al. 2008).