Recombinant human zona pellucida protein ZP3 produced by chinese hamster ovary cells induces the human sperm acrosome reaction and promotes sperm-egg fusion

• Published in: Biology of reproduction Vol. 51; no. 4; pp. 607 - 617
• Main Authors: van Duin, M, Polman, J E, De Breet, I T, van Ginneken, K, Bunschoten, H, Grootenhuis, A, Brindle, J, Aitken, R J
• Format: Journal Article
• Language: English
• Published: United States Society for the Study of Reproduction 01-10-1994
• Subjects: Acrosome - drug effects; Acrosome - physiology; Animals; Base Sequence; Blotting, Western; CHO Cells - metabolism; Chromatography, Affinity; Cricetinae; DNA, Complementary - genetics; Egg Proteins - genetics; Egg Proteins - isolation & purification; Egg Proteins - pharmacology; Female; Gene Expression; Humans; Male; Membrane Glycoproteins - genetics; Membrane Glycoproteins - isolation & purification; Membrane Glycoproteins - pharmacology; Molecular Sequence Data; Receptors, Cell Surface; Recombinant Proteins - biosynthesis; Sperm-Ovum Interactions - drug effects; Transfection; Zona Pellucida - metabolism; Zona Pellucida Glycoproteins
• ISSN: 0006-3363; 1529-7268
• DOI: 10.1095/biolreprod51.4.607

Studies on the role of specific molecules in the human fertilization process and additional assessments of potential applications
for these proteins are hampered by the limited amount of available biological material. However, this drawback might be circumvented
by the recent cloning of several gamete-specific genes, which opens possibilities for the production of recombinant proteins.
By use of cDNA and genomic DNA fragments of the human ZP3 gene, which encodes a major constituent of the zona pellucida surrounding
the oocyte, a 2.7-kb minigene was constructed containing the natural third and fourth introns of the gene and a truncated
intron between exons 2 and 3. This ZP3 DNA was transfected to Chinese hamster ovary cells, and a single-cell clone producing
the recombinant ZP3 protein (recZP3) was generated. Western blot analysis of culture medium from these cells showed that recZP3
has a molecular mass +/- 5 kDa smaller than that of natural ZP3. Under reducing conditions, it migrates at an apparent molecular
mass of 55-60 kDa. RecZP3 induced the sperm acrosome reaction and promoted fusion of human spermatozoa with zona-free hamster
oocytes, indicating that the recombinant protein is biologically active. RecZP3 provides an attractive tool for studying the
initial stage of the human fertilization process. Furthermore, it might have clinical applications in the development of diagnostic
tests for male infertility and serve as target antigen in the design of contraceptive vaccines.