Sperm surface protein PH-20 is bifunctional: One activity is a hyaluronidase and a second, distinct activity is required in secondary sperm-zona binding
Sperm surface protein PH-20 is bifunctional: One activity is a hyaluronidase and a second, distinct activity is required in secondary sperm-zona binding
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DOI：
10.1095/biolreprod55.1.80
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发表时间：
1996-07-01
影响因子：
3.6
通讯作者：
Myles, DG
中科院分区：
文献类型：
--
作者：
Hunnicutt, CR;Primakoff, P;Myles, DG
在以前的研究中，我们发现精子膜蛋白PH-20在受精的两个不同阶段起作用。对于顶体完整的精子，PH-20具有透明质酸酶活性，这是精子穿透卵母细胞周围的卵丘细胞层所必需的。在顶体反应的精子上，PH-20在精子结合（次级结合）中具有所需的功能。由于透明质酸（HA）已被检测到的透明质酸，继发精子粘附可能依赖于重复的结合和水解HA的PH-20作为透明质酸酶。或者，PH-20可以是双功能的，并且具有二级结合所需的第二种不同活性。为了区分这两种可能性，在这项研究中，我们使用了抑制PH-20在精子结合中的功能或其透明质酸酶活性的试剂。我们发现，抑制精子区结合（类似于90%）的抗PH-20单克隆抗体对透明质酸酶活性没有影响。相反，芹菜素，透明质酸酶抑制剂，阻止PH-20透明质酸酶活性93%，而不抑制精子结合。类似地，另一种抗PH-20单克隆抗体抑制透明质酸酶活性95%，仅部分抑制精子区结合（类似于45%）。我们还广泛地预处理卵母细胞与透明质酸酶，以消除所有可访问的透明质酸或透明质酸，并发现很少或没有影响次级精子区结合。我们的研究结果表明，PH-20是双功能的，有两个活动：透明质酸酶活性和第二，单独的活动所需的次级精子区结合。
In previous studies, we have found that the sperm membrane protein PH-20 acts during two different stages of fertilization. On acrosome-intact sperm, PH-20 has a hyaluronidase activity that is required for sperm penetration through the cumulus cell layer that surrounds the oocyte. On acrosome-reacted sperm, PH-20 has a required function in sperm-zona binding (secondary binding). Because hyaluronic acid (HA) has been detected in the zona pellucida, secondary sperm-zona adhesion could depend on repetitive binding and hydrolysis of HA by PH-20 acting as a hyaluronidase. Alternatively, PH-20 may be bifunctional and have a second, different activity required for secondary binding. To distinguish between these two possibilities, in this study we used reagents that inhibit either PH-20's function in sperm-zona binding or its hyaluronidase activity. We found that an anti-PH-20 monoclonal antibody that inhibited sperm-zone binding (similar to 90%) had no effect on hyaluronidase activity. Conversely, apigenin, a hyaluronidase inhibitor, blocked PH-20 hyaluronidase activity 93% without inhibiting sperm-zona binding. Similarly, another anti-PH-20 monoclonal antibody that inhibited hyaluronidase activity 95% only partially inhibited sperm-zone binding (similar to 45%). We also extensively pretreated oocytes with hyaluronidase to remove all accessible HA on or in the zona pellucida and found little or no effect on secondary sperm-zone binding. Our results suggest that PH-20 is bifunctional and has two activities: a hyaluronidase activity and a second, separate activity required for secondary sperm-zone binding.