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Recombinant human zona pellucida proteins ZP1, ZP2 and ZP3 co-expressed in a human cell line

Asian Journal of Andrology 2004年第1期6卷 3-13页

作者： MirjanaMartic EricK.Moses TimE.Adams deyiliu DebraA.Gook ClaireGarrett MarjorieE.Dunlop GordonH.W.Baker DepartmentofObstetricsandGynaecology，UniversityofMelbourne DepartmentofMedicine UniversityofMelbourne PregnancyResearchCenterRoyalWomen''sHospital ReproductiveServices RoyalWomen''sHospitalandMelbourneIVE CSIRODivisionofHealthScienceandNutrition MelbourneAustralia

Aim: To produce biologically active recombinant human (rh) ZP proteins in a human cell for use in sperm function tests. Methods: The human embryonic kidney cell line 293T was employed to produce rhZP1, rhZP2 and rhZP3 proteins individually and together by co-expression. Presence of these proteins in the culture medium and cell lysate was assessed by Western blotting analysis. The effect of the recombinant proteins on the human AR was assessed. Results: RhZP2 and rhZP3 were secreted into the culture medium, whereas rhZPl was found only in the cell lysate. Interestingly, when all zona pellucida proteins were co-expressed in the same cells, rhZPl was also secreted into the culture medium. However, despite the presence of all three ZP proteins in sufficient concentration and evidence of heavy glycosylation on gel electrophoresis, biological activity to induce the AR was not observed. Conclusion: RhZP1, rhZP2 and rhZP3 were successfully expressed in the human embryonic kidney cell line 293T. It appears that an interaction amongst these proteins may be required for release of rhZPl from the cell. Although this approach is not satisfactory for producing active human ZP proteins, it makes a significant contribution to the understanding of the structural and functional characteristics of the ZP proteins.

关键词： acrosome reaction glycosylation human cell line recombinant proteins zona pellucida

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