Interspecies diversity of zonadhesin D3-domain polypeptides in spermatozoa from animals representing five taxonomic orders. Proteins extracted from spermatozoa with Triton X-100 (TX-100) or with SDS after prior extraction with Triton X-100 (TX/SDS) were separated by SDS-PAGE, and zonadhesin was detected on Western blots with affinity-purified antibodies to the D3 domain or with antisera to the zonadhesin holoprotein. A) D3-polypeptides (protein disulfides reduced) of pig (Sus scrofa; order: Artiodactyla), bovine (Bos taurus; order: Artiodactyla), human (Homo sapiens; order: Primates), and rabbit (Oryctolagus cuniculus; order: Lagomorpha) spermatozoa detected with antibody to pig D3 domain. Note the species differences in polypeptide size, detergent solubility, or both. B) D3 polypeptides (protein disulfides reduced) of mouse, rat, and hamster spermatozoa (Mus musculus, Rattus norvegicus, and Cricetinae mesocricetus, respectively; order: Rodentia) detected with antibody to mouse D3 domain. Note the species differences in both polypeptide size and detergent solubility. C) Zonadhesin (protein disulfides not reduced) of pig, horse (Equus caballus; order: Perissodactyla), bovine, and rabbit spermatozoa detected with holoprotein antisera. Note the relatively similar size of the largest (multimeric) holoprotein form among the three ungulates.