Ascidian Sperm Acrosin and Spermosin: Structures and Roles in Fertilization
Ascidian Sperm Acrosin and Spermosin: Structures and Roles in Fertilization
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海鞘精子顶体蛋白和精子酶：结构和在受精中的作用
DOI：
10.1007/978-4-431-66982-1_9
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发表时间：
2001
期刊：
影响因子：
--
通讯作者：
H. Sawada
中科院分区：
文献类型：
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作者：
E. Kodama;T. Baba;H. Yokosawa;H. Sawada
我们以前曾报道过两种精子胰蛋白酶样蛋白酶，顶体酶和精子酶，参与海鞘Halocynthia roretzi的受精。在本研究中，我们分离的cDNA克隆编码这些酶。分离的海鞘顶体酶和精子酶前体的cDNA具有单个开放阅读框，分别编码505和388个氨基酸残基。这两种酶的mRNA在性腺中表达，但在其他组织中不表达。海鞘顶体酶具有成对的碱性残基（Lys 56-His 57），其被发现负责顶体酶（pro）与卵黄膜的结合。海鞘顶体蛋白原在C端含有两个CUB结构域，且至少有CUB结构域1参与其与卵黄膜的结合。在非还原性条件下，SDS-PAGE显示分子量为33-kDa和40-kDa的两条带，N-末端序列分析表明33-kDa的精蛋白由一条重链组成40-kDa精子素由一条重链和一条L2轻链组成（残基130-388），而40-kDa精子素由一条重链和一条L2轻链组成（残基97-129）。与L2不同，L1包含Pro富集区（L1（ΔL2））。这个富含Pro的区域似乎负责精子蛋白与卵黄外壳的结合，因为GST-L1和GST-L1（ΔL2）融合蛋白，而不是GST-L2融合蛋白，能够与28-kDa的卵黄外壳组分结合。
We have previously reported that two sperm trypsin-like proteases, acrosin and spermosin, participate in the fertilization of the ascidianHalocynthia roretzi. In the present study, we isolated the cDNA clones encoding these enzymes. The isolated cDNAs of precursors for ascidian acrosin and spermosin have single open reading frames, which encode 505 and 388 amino acid residues, respectively. The mRNAs of both enzymes are expressed in the gonads but not in other tissues. Ascidian acrosin has paired basic residues (Lys56-His57), which were found to be responsible for the binding of (pro)acrosin to the vitelline coat. It was also found that ascidian proacrosin contains two CUB domains in theC-terminal portion and that at least CUB domain 1 is involved in its binding to the vitelline coat. SDS-PAGE of the purified spermosin gave two bands with molecular masses of 33-kDa and 40-kDa under nonreducing conditions.N-terminal sequence analyses of both bands revealed that the 33-kDa spermosin is made up by a heavy chain (residues of 130–388) and an L1 light chain (97–129), while the 40-kDa spermosin consists of a heavy chain and an L2 light chain (23–129). L1, unlike L2, contains a Pro-rich region (L1(ΔL2)). This Pro-rich region appears to be responsible for the binding of spermosin to the vitelline coat, since GST-L1 and GST-L1(ΔL2) fusion proteins, but not GST-L2 fusion protein, are capable of associating with the 28-kDa vitelline coat component.