Centre for Biotechnology in Agriculture Research, Division of Genetics and Molecular Biology, Institute of Biological Sciences, Faculty of Science, University of Malaya, 50603 Kuala Lumpur, Malaysia.
Mol Biol Rep. 2012 Feb;39(2):1377-86. doi: 10.1007/s11033-011-0872-5. Epub 2011 May 26.
The prophenoloxidase activating system is an important innate immune response against microbial infections in invertebrates. The major enzyme, phenoloxidase, is synthesized as an inactive precursor and its activation to an active enzyme is mediated by a cascade of clip domain serine proteinases. In this study, a cDNA encoding a prophenoloxidase activating enzyme-III from the giant freshwater prawn Macrobrachium rosenbergii, designated as MrProAE-III, was identified and characterized. The full-length cDNA contains an open reading frame of 1110 base pair (bp) encoding a predicted protein of 370 amino acids including an 22 amino acid signal peptide. The MrProAE-III protein exhibits a characteristic sequence structure of a long serine proteases-trypsin domain and an N- and C-terminal serine proteases-trypsin family histidine active sites, respectively, which together are the characteristics of the clip-serin proteases. Sequence analysis showed that MrProAE-III exhibited the highest amino acid sequence similarity (63%) to a ProAE-III from Atlantic blue crab, Callinectes sapidus. MrProAE-III mRNA and enzyme activity of MrProAE-III were detectable in all examined tissues, including hepatopancreas, hemocytes, pleopods, walking legs, eye stalk, gill, stomach, intestine, brain and muscle with the highest level of both in hepatopancreas. This is regulated after systemic infectious hypodermal and hematopoietic necrosis virus infection supporting that it is an immune-responsive gene. These results indicate that MrProAE-III functions in the proPO system and is an important component in the prawn immune system.
酚氧化酶原激活系统是无脊椎动物抵御微生物感染的重要先天免疫反应系统。主要酶类——酚氧化酶是以无活性前体的形式合成的，其被激活为有活性的酶是由一系列 clip 结构域丝氨酸蛋白酶介导的。本研究从罗氏沼虾（Macrobrachium rosenbergii）中鉴定并表征了一个编码酚氧化酶原激活酶-III（MrProAE-III）的 cDNA。全长 cDNA 包含一个 1110 个碱基对（bp）的开放阅读框，编码一个 370 个氨基酸的预测蛋白，其中包括一个 22 个氨基酸的信号肽。MrProAE-III 蛋白表现出一个长丝氨酸蛋白酶-胰蛋白酶结构域和一个 N 端和 C 端丝氨酸蛋白酶-胰蛋白酶家族组氨酸活性位点的特征序列结构，这共同构成了 clip-丝氨酸蛋白酶的特征。序列分析表明，MrProAE-III 与大西洋蓝蟹（Callinectes sapidus）的 ProAE-III 具有最高的氨基酸序列相似性（63%）。MrProAE-III mRNA 和 MrProAE-III 酶活性可在所有检测组织中检测到，包括肝胰腺、血细胞、步足、步行足、眼柄、鳃、胃、肠、脑和肌肉，其中肝胰腺中的水平最高。这是在系统性传染性皮下和造血坏死病毒感染后进行的调节，支持它是一个免疫反应基因。这些结果表明 MrProAE-III 在原酚氧化酶系统中发挥作用，是虾类免疫系统的重要组成部分。
