BIOCHEMICAL-CHARACTERIZATION OF A GLYCOSYLPHOSPHATIDYLINOSITOL-LINKEDHYALURONIDASE ON MOUSE SPERM

On the basis of DNA homology to bee venom hyaluronidase, it was recent ly suggested that the GPI-linked mammalian sperm antigen, PH-20, may f unction as a cell surface hyaluronidase [Gmachl, M., and Kreil, G. (19 93) Proc. Natl. Acad. Sci. U.S.A. 90, 3569-3573]. We have quantified t he activity of the soluble acrosomal hyaluronidase of mouse sperm and further demonstrate the existence of a membrane-hound hyaluronidase, d etected on both acrosome-intact and acrosome-reacted mouse sperm, dist inct from the soluble form of the enzyme. The membrane-bound hyaluroni dase was specifically released by PE-PLC, indicating that it is GPI li nked. Acrosome-intact and acrosome-reacted sperm released several poly peptides (68, 44, 39, 34, 17, and 15 kDa) when treated with PI-PLC. In addition, GPI-Linked polypeptides unique to acrosome-intact or to acr osome-reacted sperm were identified. Fractionation of the PI-PLC-relea sed components from acrosome-reacted sperm using size exclusion chroma tography revealed a single peak of hyaluronidase activity which comigr ates with a 68 kDa GPI-linked protein present in these fractions. Take n together, these data demonstrate the existence of at least two isofo rms of hyaluronidase: a soluble form within the acrosomal vesicle whic h is released during acrosomal exocytosis and a GPI-linked form which is present on the surface of both acrosome-intact and acrosome-reacted sperm. Both forms may be necessary for successful penetration of the extracellular vestments that surround the egg prior to fertilization.