The mammalian egg coat participates in fertilization and prevents more than one sperm from entering the egg. Structural data pinpoint a region common to egg-coat proteins that might mediate these functions.
Eggs are surrounded by one or more extracellular coats that take part in fertilization. Mammalian eggs have a single coat, the zona pellucida, which consists of long, interconnected fibrils constructed of a few proteins1. Mouse zona-pellucida proteins are called ZP1, ZP2 and ZP3, with these last two serving as sperm receptors during fertilization1,2. All three proteins have a characteristic region known as the ZP domain3, which also occurs in related proteins that make up the coat of fish, frog and bird eggs. What's more, the ZP domain is not restricted to egg-coat proteins, but is present in hundreds of extracellular proteins found in virtually all multicellular organisms.
Mutations in the ZP domain can result in severe disorders, such as infertility, deafness and cancer. Nonetheless, no high-resolution structure of either a protein containing the ZP domain or a vertebrate protein involved in fertilization has been obtained. The propensity of ZP-domain proteins to self-assemble into polymers (through the amino-terminal half (ZP-N) of this domain4), and to undergo extensive modification after their synthesis from the gene, has made it difficult to solve their structure. But on page 653 of this issue, Monné et al.5 describe a high-resolution (2.3-ångstrom) three-dimensional structure of ZP-N from ZP3, revealing that it folds in a novel way.
The ZP domain consists of about 260 amino acids, of which eight are evolutionarily conserved cysteine residues that form disulphide linkages2. Proteins containing this domain display a mosaic architecture that is thought to arise from the combination of a common structural building block — the ZP domain — with additional sequences that have specific biological functions.
Monné and colleagues find that two-thirds of ZP-N consists of eight identical β-strands (designated A–E, E′, F and G), which are interconnected by loops of varying lengths and shapes. The strands form a sandwich consisting of two sheets, similar to that found in immunoglobulin proteins. However, the sheets rest on a platform created by the extension of strands E′, F and G (see Fig. 1 on page 654). Two conserved intramolecular disulphide bonds staple the sandwich together to stabilize the structure. The structural complexity of ZP-N meant that these features could not have been predicted from its amino-acid sequence.
The immunoglobulin superfamily includes proteins that serve as receptors, in cellular adhesion and other processes. Although ZP-N adopts an immunoglobulin-like fold, overall it has little structural similarity to other immunoglobulin-like domains and represents a new domain within this superfamily of proteins6. Nonetheless, Monné and colleagues' data5 indicate that the platform region of the molecule participates in intermolecular interactions. This region contains two conserved tyrosine residues that interact with each other. When one of the two tyrosines is mutated7, or when either of them is missing8, ZP-domain proteins fail to form polymers. Collectively, these observations strongly suggest that the unique fold in ZP-N triggers the polymerization of ZP-domain proteins into higher-order structures. In the zona pellucida, for instance, this region might mediate the polymerization of the ZP2 and ZP3 dimers that form the long fibrils interconnected by ZP1 (ref. 2).
Monné et al. also present a structural model that takes into account the proposal9 that egg-coat proteins such as ZP1 and ZP2 in frogs, birds and mammals have several copies of ZP-N upstream of the ZP domain (Fig. 1). Only ZP-domain proteins involved in fertilization contain the extra copies of ZP-N, and in ZP2, these form extended, rod-like structures. It is possible that interaction between these structures on different ZP2 molecules accounts for the post-fertilization compaction of zona-pellucida fibrils that creates a barrier to further sperm penetration and masks sperm-binding sites on ZP-N. The copies of ZP-N have an unusually high degree of amino-acid diversity among different species. Amino-acid variation also occurs in egg-coat proteins that mediate sperm recognition in both vertebrates and invertebrates, and this variation is thought to be important for speciation10,11. So perhaps amino-acid diversity in the ZP-N repeats of ZP2 are necessary for species-restricted fertilization in mammals.
Monné and colleagues' observations provide intriguing insights into the polymerization of ZP-domain proteins in general and aspects of mammalian fertilization in particular. A full understanding of the relationships between ZP2 and ZP3 in zona-pellucida fibrils, however, will require further structural studies of the entire ZP domain. For example, it is unclear why both ZP2 and ZP3 are required for the zona pellucida to assemble around the egg2; certainly, some ZP-domain-containing proteins, such as uromodulin, can polymerize into fibrils and form a matrix on their own. Perhaps the carboxy-terminal region of the ZP domain of ZP2 and ZP3 imparts specificity to interactions between the proteins. The structure of an entire ZP domain could shed light on this and other issues.
Determining high-resolution structures of ZP2 and ZP3 should be most illuminating. In the case of ZP3, the polypeptide downstream of the ZP domain serves as an initial binding site for sperm, whereas the polypeptide in the ZP domain of ZP2 — including the extra copies of ZP-N — provides subsequent binding sites for sperm. So stay tuned for the exciting instalments that are sure to follow this tantalizing first glimpse of zona-pellucida protein structures5.
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About this article
Heterozygous mutations in ZP1 and ZP3 cause formation disorder of ZP and female infertility in human
Journal of Cellular and Molecular Medicine (2020)
Annual Review of Genomics and Human Genetics (2012)