title: Antimicrobial actions of human and macaque sperm associated antigen (SPAG) 11 isoforms: influence of the N-terminal peptide.
abstract: In addition to their role in sperm maturation, recent evidence has indicated that epididymal proteins have a role in male reproductive tract innate immunity. Herein we demonstrate that human and macaque epididymal protein isoforms in the SPAG (sperm associated antigen) 11 family, full length SPAG11C, K and L exhibit potent antibacterial activity against E. coli. Analysis of activities of the N- and C-terminal domains revealed that the human N-terminal peptide is bactericidal, while the C-terminal domains that contain the defensin-like 6 cysteine array in SPAG11C and partial arrays in SPAG11K and SPAG11L, lack antibacterial activity. The N-terminal peptide does not appear to contain all the determinants of activity since full-length human SPAG11C is more active than the isolated N-terminal peptide and since sulfhydryl reduction and alkylation, which would affect primarily the C-terminal peptides, completely abolished activities of the whole proteins. These results suggest that the structure conferred by the disulfide bonds in human SPAG11C contributes to the antibacterial activity of the whole molecule. The activities of the N-terminal peptide and of full length human SPAG11C were somewhat reduced in increasing NaCl concentrations. In contrast, the antibacterial activities of full length macaque SPAG11C, K and L were unaffected by the presence of NaCl suggesting a mechanism in the macaque that is less dependent upon electrostatic interactions. SPAG11C, K and L disrupted E. coli membranes but had no effect on erythrocyte membranes. Inhibition of E. coli RNA, DNA and protein synthesis by nonlethal concentrations of SPAG11 isoforms indicated an additional mechanism of bacterial killing.
authors.affiliation: Laboratories for Reproductive Biology, Department of Pediatrics, University of North Carolina, Chapel Hill, 27599-7500, USA.
authors.forename: Suresh
authors.initials: S
authors.lastname: Yenugu
authors.forename: Katherine G
authors.initials: KG
authors.lastname: Hamil
authors.forename: Frank S
authors.initials: FS
authors.lastname: French
authors.forename: Susan H
authors.initials: SH
authors.lastname: Hall
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D000954:Antigens, Surface / Q000494:pharmacology / Q000502:physiology*; D002462:Cell Membrane / Q000187:drug effects / Q000648:ultrastructure; D002463:Cell Membrane Permeability; D004926:Escherichia coli / Q000187:drug effects / Q000378:metabolism; D005456:Fluorescent Dyes; D006020:Glycopeptides / Q000494:pharmacology / Q000502:physiology*; D006461:Hemolysis; D006801:Humans; D066298:In Vitro Techniques; D008251:Macaca; D008297:Male; D008855:Microscopy, Electron, Scanning; D008969:Molecular Sequence Data; D010455:Peptides / Q000494:pharmacology*; D020033:Protein Isoforms / Q000494:pharmacology / Q000502:physiology; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000494:pharmacology
chemical_list: D000900:Anti-Bacterial Agents; D000954:Antigens, Surface; D005456:Fluorescent Dyes; D006020:Glycopeptides; D010455:Peptides; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; C087044:SPAG11A protein, human