Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 1998 Mar 1;330(Pt 2):839–845. doi: 10.1042/bj3300839

The polypeptide backbone of recombinant human zona pellucida glycoprotein-3 initiates acrosomal exocytosis in human spermatozoa in vitro.

N Chapman 1, E Kessopoulou 1, P Andrews 1, D Hornby 1, C R Barratt 1
PMCID: PMC1219214  PMID: 9480899

Abstract

Human gamete interaction is of fundamental biological importance, yet the molecular interactions between spermatozoa and the zona pellucida are poorly understood. Surprisingly, the role of the polypeptide backbone of zona pellucida glycoprotein 3 (ZP3), the putative ligand for spermatozoa activation, has been largely overlooked. Purified recombinant human ZP3 was expressed in Escherichia coli as a C-terminal fusion to the dimeric glutathione S-transferase (GST) from Schistosoma japonicum and was shown to induce acrosomal exocytosis in live, capacitated human spermatozoa. The level of exocytosis is comparable with that obtained using purified, glycosylated, recombinant human ZP3 [van Duin, M., Polman, J.E.M., DeBreet, I.T.M., Van Ginneken, K., Bunschoten, H., Grootenhuis, A., Brindle, J. and Aitken, R.J. (1994). Biol Reprod. 51, 607-617]. These data imply that the polypeptide chain of human ZP3 contributes to recognition of spermatozoa during acrosomal exocytosis in vitro.

Full Text

The Full Text of this article is available as a PDF (339.4 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bagavant H., Yurewicz E. C., Sacco A. G., Talwar G. P., Gupta S. K. Block in porcine gamete interaction by polyclonal antibodies to a pig ZP3 beta fragment having partial sequence homology to human ZP3. J Reprod Immunol. 1993 Dec;25(3):277–283. doi: 10.1016/0165-0378(93)90069-t. [DOI] [PubMed] [Google Scholar]
  2. Beebe S. J., Leyton L., Burks D., Ishikawa M., Fuerst T., Dean J., Saling P. Recombinant mouse ZP3 inhibits sperm binding and induces the acrosome reaction. Dev Biol. 1992 May;151(1):48–54. doi: 10.1016/0012-1606(92)90212-y. [DOI] [PubMed] [Google Scholar]
  3. Bleil J. D., Greve J. M., Wassarman P. M. Identification of a secondary sperm receptor in the mouse egg zona pellucida: role in maintenance of binding of acrosome-reacted sperm to eggs. Dev Biol. 1988 Aug;128(2):376–385. doi: 10.1016/0012-1606(88)90299-0. [DOI] [PubMed] [Google Scholar]
  4. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  5. Chamberlin M. E., Dean J. Human homolog of the mouse sperm receptor. Proc Natl Acad Sci U S A. 1990 Aug;87(16):6014–6018. doi: 10.1073/pnas.87.16.6014. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Clements S., Cooke I. D., Barratt C. L. Implementing comprehensive quality control in the andrology laboratory. Hum Reprod. 1995 Aug;10(8):2096–2106. doi: 10.1093/oxfordjournals.humrep.a136242. [DOI] [PubMed] [Google Scholar]
  7. Cross N. L., Morales P., Overstreet J. W., Hanson F. W. Induction of acrosome reactions by the human zona pellucida. Biol Reprod. 1988 Feb;38(1):235–244. doi: 10.1095/biolreprod38.1.235. [DOI] [PubMed] [Google Scholar]
  8. Florman H. M., Bechtol K. B., Wassarman P. M. Enzymatic dissection of the functions of the mouse egg's receptor for sperm. Dev Biol. 1984 Nov;106(1):243–255. doi: 10.1016/0012-1606(84)90079-4. [DOI] [PubMed] [Google Scholar]
  9. Florman H. M., Wassarman P. M. O-linked oligosaccharides of mouse egg ZP3 account for its sperm receptor activity. Cell. 1985 May;41(1):313–324. doi: 10.1016/0092-8674(85)90084-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Franken D. R., Morales P. J., Habenicht U. F. Inhibition of G protein in human sperm and its influence on acrosome reaction and zona pellucida binding. Fertil Steril. 1996 Dec;66(6):1009–1011. [PubMed] [Google Scholar]
  11. Grant S. G., Jessee J., Bloom F. R., Hanahan D. Differential plasmid rescue from transgenic mouse DNAs into Escherichia coli methylation-restriction mutants. Proc Natl Acad Sci U S A. 1990 Jun;87(12):4645–4649. doi: 10.1073/pnas.87.12.4645. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Guan K. L., Dixon J. E. Eukaryotic proteins expressed in Escherichia coli: an improved thrombin cleavage and purification procedure of fusion proteins with glutathione S-transferase. Anal Biochem. 1991 Feb 1;192(2):262–267. doi: 10.1016/0003-2697(91)90534-z. [DOI] [PubMed] [Google Scholar]
  13. Harris J. D., Hibler D. W., Fontenot G. K., Hsu K. T., Yurewicz E. C., Sacco A. G. Cloning and characterization of zona pellucida genes and cDNAs from a variety of mammalian species: the ZPA, ZPB and ZPC gene families. DNA Seq. 1994;4(6):361–393. doi: 10.3109/10425179409010186. [DOI] [PubMed] [Google Scholar]
  14. Henderson R. M., Schneider S., Li Q., Hornby D., White S. J., Oberleithner H. Imaging ROMK1 inwardly rectifying ATP-sensitive K+ channel protein using atomic force microscopy. Proc Natl Acad Sci U S A. 1996 Aug 6;93(16):8756–8760. doi: 10.1073/pnas.93.16.8756. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Karlsson K. A. Microbial recognition of target-cell glycoconjugates. Curr Opin Struct Biol. 1995 Oct;5(5):622–635. doi: 10.1016/0959-440x(95)80054-9. [DOI] [PubMed] [Google Scholar]
  16. Lasky L. A. Selectin-carbohydrate interactions and the initiation of the inflammatory response. Annu Rev Biochem. 1995;64:113–139. doi: 10.1146/annurev.bi.64.070195.000553. [DOI] [PubMed] [Google Scholar]
  17. Leyton L., Saling P. Evidence that aggregation of mouse sperm receptors by ZP3 triggers the acrosome reaction. J Cell Biol. 1989 Jun;108(6):2163–2168. doi: 10.1083/jcb.108.6.2163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Liu D. Y., Baker H. W. A simple method for assessment of the human acrosome reaction of spermatozoa bound to the zona pellucida: lack of relationship with ionophore A23187-induced acrosome reaction. Hum Reprod. 1996 Mar;11(3):551–557. doi: 10.1093/humrep/11.3.551. [DOI] [PubMed] [Google Scholar]
  19. Macek M. B., Lopez L. C., Shur B. D. Aggregation of beta-1,4-galactosyltransferase on mouse sperm induces the acrosome reaction. Dev Biol. 1991 Oct;147(2):440–444. doi: 10.1016/0012-1606(91)90301-i. [DOI] [PubMed] [Google Scholar]
  20. McTigue M. A., Williams D. R., Tainer J. A. Crystal structures of a schistosomal drug and vaccine target: glutathione S-transferase from Schistosoma japonica and its complex with the leading antischistosomal drug praziquantel. J Mol Biol. 1995 Feb 10;246(1):21–27. doi: 10.1006/jmbi.1994.0061. [DOI] [PubMed] [Google Scholar]
  21. Millar S. E., Chamow S. M., Baur A. W., Oliver C., Robey F., Dean J. Vaccination with a synthetic zona pellucida peptide produces long-term contraception in female mice. Science. 1989 Nov 17;246(4932):935–938. doi: 10.1126/science.2479101. [DOI] [PubMed] [Google Scholar]
  22. Miska W., Fehl P., Henkel R. Biochemical and immunological characterization of the acrosome reaction-inducing substance (ARIS) of hFF. Biochem Biophys Res Commun. 1994 Feb 28;199(1):125–129. doi: 10.1006/bbrc.1994.1203. [DOI] [PubMed] [Google Scholar]
  23. Moore H. D., Smith C. A., Hartman T. D., Bye A. P. Visualization and characterization of the acrosome reaction of human spermatozoa by immunolocalization with monoclonal antibody. Gamete Res. 1987 Jul;17(3):245–249. doi: 10.1002/mrd.1120170308. [DOI] [PubMed] [Google Scholar]
  24. Murase T., Roldan E. R. Epidermal growth factor stimulates hydrolysis of phosphatidylinositol 4,5-bisphosphate, generation of diacylglycerol and exocytosis in mouse spermatozoa. FEBS Lett. 1995 Mar 6;360(3):242–246. doi: 10.1016/0014-5793(95)00114-o. [DOI] [PubMed] [Google Scholar]
  25. Nakano H., Yamazaki T., Ikeda M., Masai H., Miyatake S., Saito T. Purification of glutathione S-transferase fusion proteins as a non-degraded form by using a protease-negative E. coli strain, AD202. Nucleic Acids Res. 1994 Feb 11;22(3):543–544. doi: 10.1093/nar/22.3.543. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Oehninger S., Hinsch E., Pfisterer S., Veeck L. L., Kolm P., Schill W. B., Hodgen G. D., Hinsch K. D. Use of a specific zona pellucida (ZP) protein 3 antiserum as a clinical marker for human ZP integrity and function. Fertil Steril. 1996 Jan;65(1):139–145. doi: 10.1016/s0015-0282(16)58041-8. [DOI] [PubMed] [Google Scholar]
  27. Ohlendieck K., Lennarz W. J. Role of the sea urchin egg receptor for sperm in gamete interactions. Trends Biochem Sci. 1995 Jan;20(1):29–33. doi: 10.1016/s0968-0004(00)88947-1. [DOI] [PubMed] [Google Scholar]
  28. Perry R. L., Naeeni M., Barratt C. L., Warren M. A., Cooke I. D. A time course study of capacitation and the acrosome reaction in human spermatozoa using a revised chlortetracycline pattern classification. Fertil Steril. 1995 Jul;64(1):150–159. [PubMed] [Google Scholar]
  29. Pollitt S., Zalkin H. Role of primary structure and disulfide bond formation in beta-lactamase secretion. J Bacteriol. 1983 Jan;153(1):27–32. doi: 10.1128/jb.153.1.27-32.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Preibisch G., Ishihara H., Tripier D., Leineweber M. Unexpected translation initiation within the coding region of eukaryotic genes expressed in Escherichia coli. Gene. 1988 Dec 10;72(1-2):179–186. doi: 10.1016/0378-1119(88)90141-2. [DOI] [PubMed] [Google Scholar]
  31. Rickert K. W., Imperiali B. Analysis of the conserved glycosylation site in the nicotinic acetylcholine receptor: potential roles in complex assembly. Chem Biol. 1995 Nov;2(11):751–759. doi: 10.1016/1074-5521(95)90103-5. [DOI] [PubMed] [Google Scholar]
  32. Sacco A. G., Yurewicz E. C., Subramanian M. G., Matzat P. D. Porcine zona pellucida: association of sperm receptor activity with the alpha-glycoprotein component of the Mr = 55,000 family. Biol Reprod. 1989 Sep;41(3):523–532. doi: 10.1095/biolreprod41.3.523. [DOI] [PubMed] [Google Scholar]
  33. Sacco A. G., Yurewicz E. C., Subraminian M. G., DeMayo F. J. Zona pellucida composition: species cross reactivity and contraceptive potential of antiserum to a purified pig zona antigen (PPZA). Biol Reprod. 1981 Dec;25(5):997–1008. doi: 10.1095/biolreprod25.5.997. [DOI] [PubMed] [Google Scholar]
  34. Saragüeta P., Lanuza G., Miranda P. V., Tezón J. G., Barañao J. L. Immunoglobulins from human follicular fluid induce the acrosome reaction in human sperm. Mol Reprod Dev. 1994 Nov;39(3):280–288. doi: 10.1002/mrd.1080390305. [DOI] [PubMed] [Google Scholar]
  35. Sareneva T., Pirhonen J., Cantell K., Kalkkinen N., Julkunen I. Role of N-glycosylation in the synthesis, dimerization and secretion of human interferon-gamma. Biochem J. 1994 Nov 1;303(Pt 3):831–840. doi: 10.1042/bj3030831. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Sherrill J. M., Kyte J. Activation of epidermal growth factor receptor by epidermal growth factor. Biochemistry. 1996 May 7;35(18):5705–5718. doi: 10.1021/bi9602268. [DOI] [PubMed] [Google Scholar]
  37. Shine J., Dalgarno L. The 3'-terminal sequence of Escherichia coli 16S ribosomal RNA: complementarity to nonsense triplets and ribosome binding sites. Proc Natl Acad Sci U S A. 1974 Apr;71(4):1342–1346. doi: 10.1073/pnas.71.4.1342. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Vandevoort C. A., Cherr G. N., Overstreet J. W. Hyaluronic acid enhances the zona pellucida-induced acrosome reaction of macaque sperm. J Androl. 1997 Jan-Feb;18(1):1–5. [PubMed] [Google Scholar]
  39. Wang C., Eufemi M., Turano C., Giartosio A. Influence of the carbohydrate moiety on the stability of glycoproteins. Biochemistry. 1996 Jun 11;35(23):7299–7307. doi: 10.1021/bi9517704. [DOI] [PubMed] [Google Scholar]
  40. Wassarman P. M. Profile of a mammalian sperm receptor. Development. 1990 Jan;108(1):1–17. doi: 10.1242/dev.108.Supplement.1. [DOI] [PubMed] [Google Scholar]
  41. Wells J. A. Binding in the growth hormone receptor complex. Proc Natl Acad Sci U S A. 1996 Jan 9;93(1):1–6. doi: 10.1073/pnas.93.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Whitmarsh A. J., Woolnough M. J., Moore H. D., Hornby D. P., Barratt C. L. Biological activity of recombinant human ZP3 produced in vitro: potential for a sperm function test. Mol Hum Reprod. 1996 Dec;2(12):911–919. doi: 10.1093/molehr/2.12.911. [DOI] [PubMed] [Google Scholar]
  43. van Duin M., Polman J. E., De Breet I. T., van Ginneken K., Bunschoten H., Grootenhuis A., Brindle J., Aitken R. J. Recombinant human zona pellucida protein ZP3 produced by chinese hamster ovary cells induces the human sperm acrosome reaction and promotes sperm-egg fusion. Biol Reprod. 1994 Oct;51(4):607–617. doi: 10.1095/biolreprod51.4.607. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES