Horseshoe crab sperm contain a unique isoform of arginine kinase that is present in midpiece and flagellum

Spermatozoa of the horseshoe crab, Limulus polyphemus, contained high activities of arginine kinase (AK), the bulk of which was completely solubilized in buffer lacking detergent. Sperm AK, along with the fast isoform of muscle AK, were purified to homogeneity. Sperm AK had similar electrophoretic mobility on cellulose acetate to that of muscle AK. In contrast, sperm AK ran somewhat slower than muscle AK on SDS‐PAGE, indicating that this protein has a higher relative molecular mass (44 kD vs. 41 kD). These two forms also showed different chromatographic behavior on a reverse‐phase HPLC column. Peptide maps of cyanogen bromide cleavage fragments of sperm and muscle AK showed distinct differences. Collectively, these data suggest that the sperm of L. polyphemus contain a unique AK isoform. Polyclonal anti‐muscle AK antibodies, which showed strong reactivity against sperm AK, were used to localize AK in sperm. Both indirect immunofluorescence and immunogold transmission electron microscopy showed that AK was present in the midpiece in the vicinity of the mitochondria and also along the length of the flagellum. Given the polarized geometry of these cells, it appears that the AK reaction may play a role in high energy phosphate transport to dynein ATPases in the flagellum.