Prophenoloxidase-activating proteinase-3 (PAP-3) from Manduca sexta hemolymph:: a clip-domain serine proteinase regulated by serpin-1J and serine proteinase homologs
Prophenoloxidase-activating proteinase-3 (PAP-3) from Manduca sexta hemolymph:: a clip-domain serine proteinase regulated by serpin-1J and serine proteinase homologs
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DOI：
10.1016/s0965-1748(03)00123-1
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发表时间：
2003-10-01
影响因子：
3.8
通讯作者：
Kanost, M
中科院分区：
文献类型：
--
作者：
Jiang, HB;Wang, Y;Kanost, M
酚氧化酶 (PO) 是一种关键酶，与昆虫和甲壳类动物的多种防御机制有关。它是由原酚氧化酶 (proPO) 通过原酚氧化酶激活蛋白酶 (PAP) 的有限蛋白水解而转化而来。我们之前从烟草天蛾 Manduca sexta 的珠被中分离出 PAP-1，从血淋巴中分离出 PAP-2。在这里，我们报告纯化、表征。以及血淋巴对 PAP-3 的调节。与 M. sexta PAP-2 类似，PAP-3 由两个氨基末端夹结构域和一个羧基末端催化结构域组成，而 PAP-1 在其氨基末端仅包含一个夹结构域。纯化的 PAP-3 在 Arg(51) 处裂解 proPO，并产生低水平的 PO 活性。然而，当 M. sexta 丝氨酸蛋白酶同源物-1 和 -2 存在时，该酶有效地激活 proPO。这些蛋白酶样蛋白与免疫凝集素 2（脂多糖的模式识别受体）相关。 M. sexta PAP-3 被重组 serpin-1J 抑制，重组 serpin-1J 与该酶形成 SDS 稳定的复合物。在幼虫的脂肪体或血细胞中检测到 PAP-3 mRNA 水平较低，但在受到细菌攻击的昆虫中水平升高。这些数据以及我们之前对 PAP-1 和 PAP-2 的结果表明，PAP 激活 proPO 是一个严格调控的过程。个体 PAP 在免疫反应和发育过程中可以发挥不同的作用。 (C) 2003 Elsevier Ltd. 保留所有权利。
Phenoloxidase (PO) is a key enzyme implicated in several defense mechanisms in insects and crustaceans. It is converted from prophenoloxidase (proPO) through limited proteolysis by prophenoloxidase-activating proteinase (PAP). We previously isolated PAP-1 from integument and PAP-2 from hemolymph of the tobacco hornworm, Manduca sexta. Here, we report the purification, characterization. and regulation of PAP-3 from the hemolymph. Similar to M. sexta PAP-2, PAP-3 consists of two amino-terminal clip domains followed by a carboxyl-terminal catalytic domain, whereas PAP-1 contains only one clip domain at its amino-terminus. Purified PAP-3 cleaved proPO at Arg(51) and generated a low level of PO activity. However, the enzyme efficiently activated proPO when M. sexta serine proteinase homolog-1 and -2 were present. These proteinase-like proteins associate with immulectin-2, a pattern-recognition receptor for lipopolysaccharide. M. sexta PAP-3 was inhibited by recombinant serpin-1J, which formed an SDS-stable complex with the enzyme. PAP-3 mRNA was detected at a low level in the fat body or hemocytes of naive larvae, but was elevated in insects that had been challenged with bacteria. These data, along with our previous results on PAP-1 and PAP-2, indicate that proPO activation by PAPs is a tightly regulated process. Individual PAPs could play different roles during immune responses and developmental processes. (C) 2003 Elsevier Ltd. All rights reserved.