Molecular determinants of Xolloid action in vivo
Molecular determinants of Xolloid action in vivo
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DOI：
10.1074/jbc.m804232200
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发表时间：
2008-10-03
影响因子：
4.8
通讯作者：
Dale, Leslie
中科院分区：
文献类型：
--
作者：
Geach, Timothy J.;Dale, Leslie
XLD(Xolloid)是非洲爪蛙胚胎中发现的Tolloid金属蛋白水解酶家族的成员。它切割BMP2/4的抑制性结合蛋白Chordin，释放与这些重要的腹壁信号亲和力降低的片段。因此，XLD活性的增加会使非洲爪哇胚胎腹侧。我们已经使用这种表型作为一种分析方法来确定XLD的C末端非蛋白水解酶成分对体内活性的需求。这部分蛋白由5个补体C1r/C1s-海胆表皮生长因子-BMP1(CUB)和两个表皮生长因子结构域组成，它们被认为参与蛋白质之间的相互作用，并可能赋予底物特异性。我们的结果表明，与CUB1和CUB2偶联的蛋白酶是非洲爪哇胚胎腹侧化和阻断Chordin的背轴诱导活性所需的最小结构域。XLD-CUB1-CUB2切割Chordin，而蛋白酶失活版本共沉淀Chordin。我们的结果表明，第一个和第二个CUB结构域与Chordin结合，并将其呈现给蛋白酶结构域。非蛋白水解酶XLD阻断野生型XLD对Chordin的切割，并使注入的非洲爪哇胚胎背部。我们发现蛋白酶失活的XLD-CUB1-CUB2不具有这一活性，并且所有的C末端结构域都是产生背化表型所必需的。
Xld (Xolloid) is a member of the Tolloid family of metalloproteases found in embryos of the frog Xenopus laevis. It cleaves Chordin, an inhibitory binding protein for BMP2/4, releasing fragments with reduced affinity for these important ventralizing signals. As a consequence, increasing Xld activity ventralizes Xenopus embryos. We have used this phenotype as an assay to determine the requirement for the C-terminal, nonprotease component of Xld for in vivo activity. This part of the protein is composed of five complement C1r/C1s-sea urchin epidermal growth factor-BMP1 (CUB) and two epidermal growth factor domains, which are thought to be involved in protein-protein interactions and may confer substrate specificity. Our results show that the protease coupled to CUB1 and CUB2 is the minimum domain structure required to ventralize Xenopus embryos and to block the dorsal axis-inducing activity of Chordin. Xld-CUB1-CUB2 cleaves Chordin, and a protease-inactive version co-precipitates Chordin. Our results indicate that the first and second CUB domains bind Chordin and present it to the protease domain. Protease-inactive Xld blocks the cleavage of Chordin by wild-type Xld and dorsalizes injected Xenopus embryos. We find that protease-inactive Xld-CUB1-CUB2 does not share this activity and that all of the C-terminal domains are required to generate the dorsalized phenotype.