The zona pellucida is an extracellular matrix consisting of three
glycoproteins that surrounds mammalian eggs and mediates fertilization. The primary structures of mouse ZP1, ZP2, and ZP3 have been deduced from
cDNA. Each has a predicted
signal peptide and a transmembrane domain from which an ectodomain must be released. All three
zona proteins undergo extensive co- and post-translational modifications important for secretion and assembly of the
zona matrix. In this report, native zonae pellucidae were isolated and structural features of individual
zona proteins within the mixture were determined by high resolution electrospray mass spectrometry. Complete coverage of the primary structure of native ZP3, 96% of ZP2, and 56% of ZP1, the least abundant
zona protein, was obtained. Partial
disulfide bond assignments were made for each
zona protein, and the size of the processed, native
protein was determined. The N termini of ZP1 and ZP3, but not ZP2, were blocked by cyclization of
glutamine to
pyroglutamate. The C termini of ZP1, ZP2, and ZP3 lie upstream of a dibasic motif, which is part of, but distinct from, a
proprotein convertase cleavage site. The
zona proteins are highly glycosylated and 4/4 potential N-linkage sites on ZP1, 6/6 on ZP2, and 5/6 on ZP3 are occupied. Potential O-linked
carbohydrate sites are more ubiquitous, but less utilized.