Sperm surface protein PH-20 is bifunctional: One activity is a hyaluronidase and a second, distinct activity is required in secondary sperm-zona binding
Hunnicutt,Gary R.; Primakoff,Paul; Myles,Diane G.
Biology of Reproduction 55(1): 80-86
Publication date: 1996
In previous studies, we have found that the sperm membrane protein PH- 20acts during two different stages of fertilization. On acrosome- intactsperm, PH-20 has a hyaluronidase activity that is required for spermpenetration through the cumulus cell layer that surrounds the oocyte. Onacrosome-reacted sperm, PH-20 has a required function in sperm-zona binding(secondary binding). Because hyaluronic acid (HA) has been detected in thezona pellucida, secondary sperm-zona adhesion could depend on repetitivebinding and hydrolysis of HA by PH-20 acting as a hyaluronidase.Alternatively, PH-20 may be bifunctional and have a second, differentactivity required for secondary binding. To distinguish between these twopossibilities, in this study we used reagents that inhibit either PH-20'sfunction in sperm-zona binding or its hyaluronidase activity. We found thatan anti-PH-20 monoclonal antibody that inhibited sperm-zona binding(approximately 90%) had no effect on hyaluronidase activity. Conversely,apigenin, a hyaluronidase inhibitor, blocked PH-20 hyaluronidase activity93% without inhibiting sperm-zona binding. Similarly, another anti-PH-20monoclonal antibody that inhibited hyaluronidase activity 95% onlypartially inhibited sperm-zona binding (approximately 45%). We alsoextensively pretreated oocytes with hyaluronidase to remove all accessibleHA on or in the zona pellucida and found little or no effect on secondarysperm-zona binding. Our results suggest that PH-20 is bifunctional and hastwo activities: a hyaluronidase activity and a second, separate activityrequired for secondary sperm-zona binding.