EVIDENCE FOR ACROSIN-LIKE ENZYME IN SPERM EXTRACT AND ITS INVOLVEMENT IN FERTILIZATION OF THE ASCIDIAN, HALOCYNTHIA RORETZI
EVIDENCE FOR ACROSIN-LIKE ENZYME IN SPERM EXTRACT AND ITS INVOLVEMENT IN FERTILIZATION OF THE ASCIDIAN, HALOCYNTHIA RORETZI
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DOI：
10.1002/mrd.1120050309
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发表时间：
1982-01-01
期刊：
影响因子：
--
通讯作者：
ISHII, S
中科院分区：
文献类型：
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作者：
SAWADA, H;YOKOSAWA, H;ISHII, S
用t-butyloxycarbonyl-L-Val-L-Pro-L-Arg-4-methylcoumaryl-7-amide(Boc Val-Pro-Arg-MCA)和其他精氨酸基或赖氨基MCA衍生物作为底物，证明了罗氏单胞藻的精子中存在一种蛋白酶。在粗提物中对该酶的几种性质进行了研究。该酶的最适pH值在8.0附近，CaCl2的加入对酶活性有一定的促进作用。在最佳条件下测定了Boc-Val-Pro-Arg-MCA的Km值为87µm。凝胶过滤的表观相对分子质量估计为35,000。该酶可被氟磷酸二异丙酯、亮蛋白、止痛剂、对氨基苯甲酰胺、Val-Pro-Arg-CH2Cl和大豆胰酶抑制剂抑制，而对糜抑素、弹性蛋白、对氯汞苯甲酸、对甲苯磺酰-赖氨酸-甲苯磺酰-苯丙氨酸-甲苯几乎不抑制。BOC-Val-Pro-Arg-MCA是最敏感的底物，对海囊卵受精的抑制效果最好。海鞘精子提取液中的这种酶具有与哺乳动物顶体酶非常相似的特性[EC 3.4.21.10]，该酶作为一种溶酶参与受精。
The presence of a protease was demonstrated in sperm of the solitary ascidian, H. roretzi, by using t-butyloxycarbonyl-L-Val-L-Pro-L-Arg-4-methylcoumaryl-7-amide (Boc Val-Pro-Arg-MCA) and other arginyl or lysyl MCA derivatives as substrates. Several properties of the enzyme were investigated in a crude extract. The activity had a pH optimum near 8.0 and was enhanced by the addition of CaCl2. The Km value of 87 .mu.M was determined for Boc-Val-Pro-Arg-MCA under the optimal conditions. An apparent MW of 35,000 by gel filtration was estimated. The enzyme was inhibited with diisopropyl fluorophosphate, leupeptin, antipain, p-aminobenzamidine, Val-Pro-Arg-CH2Cl and soybean trypsin inhibitor, but scarcely inhibited with chymostatin, elastatinal, p-chloromercuribenzoic acid, tosyl-Lys-CH2Cl and tosyl-Phe-CH2Cl. Boc-Val-Pro-Arg-MCA, the most susceptible of the substrates examined, showed the most effective inhibition against fertilization of ascidian eggs. This enzyme in ascidian sperm extract has features closely similar to mammalian acrosin [EC 3.4.21.10], and the enzyme is involved in fertilization as one of the lysins.