title: High-affinity binding of seminal plasma PSP94 to human immunoglobulin is through the Fab domain.
abstract: Prostate secretory protein of 94 amino acids (PSP94) is one of the major proteins present in human seminal plasma. We had earlier reported that PSP94 has the ability to bind to human IgG. The aims of the present study were to further delineate the PSP94-IgG interaction and to understand whether this could have any significance in sperm function. Direct binding of IgG fragments to PSP94 showed maximal binding with F(ab')(2) followed by Fab, while Fc displayed least binding in ELISA. Binding kinetics of PSP94-IgG interaction using surface plasmon resonance (SPR) revealed high-affinity binding of IgG to PSP94 with a dissociation constant (K(D)) of 8.8 x 10(-)(11)M. PSP94-IgG interaction was found to be through the Fab domains of IgG. Real-time interaction kinetics revealed association constants for binding of IgG, Fab, and F(ab')(2) towards PSP94 to be of the same order but with altered dissociation constants. IgG and its F(ab')(2) fragment once complexed to PSP94 demonstrated negligible dissociation, while dissociation rate of Fab fragment was 6.6 x 10(-)(4). In silico molecular modeling of PSP94-IgG complex identified N- and C-terminal beta-strands of PSP94 to be the most plausible region involved in IgG interaction. Immunofluorescence studies revealed that IgG bound to human spermatozoa predominantly in the tail region, which could be prevented when IgG was preincubated with PSP94. This study reports for the first time that IgG forms a high-affinity complex with PSP94 through its F(ab')(2) domain and reveals the ability of PSP94 to prevent binding of IgG to spermatozoa.
authors.affiliation: Division of Structural Biology and Biomedical Informatics Centre, National Institute for Research in Reproductive Health (ICMR), Jehangir Merwanji Street, Parel, Mumbai-400012, India.
authors.forename: Dhanashree D
authors.initials: DD
authors.lastname: Jagtap
authors.forename: Deepak N
authors.initials: DN
authors.lastname: Modi
authors.forename: Mukesh
authors.initials: M
authors.lastname: Kumar
authors.forename: Bhakti R
authors.initials: BR
authors.lastname: Pathak
authors.forename: Smita D
authors.initials: SD
authors.lastname: Mahale
mesh_terms: D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000378:metabolism*; D007074:Immunoglobulin G / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008958:Models, Molecular; D029584:Prostatic Secretory Proteins / Q000378:metabolism*; D011487:Protein Conformation; D012661:Semen / Q000378:metabolism*; D013094:Spermatozoa / Q000378:metabolism*; D020349:Surface Plasmon Resonance
chemical_list: D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D029584:Prostatic Secretory Proteins; C043076:beta-microseminoprotein