Pepsin-like aspartic protease (Sc-ASP155) cloning, molecular characterization and gene expression analysis in developmental stages of nematode Steinernema carpocapsae
Pepsin-like aspartic protease (Sc-ASP155) cloning, molecular characterization and gene expression analysis in developmental stages of nematode Steinernema carpocapsae
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DOI：
10.1016/j.gene.2012.03.062
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发表时间：
2012-06-01
期刊：
影响因子：
3.5
通讯作者：
Simoes, Nelson
中科院分区：
文献类型：
--
作者：
Balasubramanian, Natesan;Nascimento, Gisela;Simoes, Nelson
Steinernema carpocapsae是一种昆虫寄生线虫，与细菌嗜热小杆菌（Xellobdus cereophila）相关。这些共生复合物对昆虫宿主是致命的。许多蛋白酶基因先前被证明在寄生期间被诱导，包括预测编码天冬氨酸蛋白酶的基因，该基因在本研究中被克隆和分析。根据EST片段克隆了编码Sc-ASP 155的cDNA。Sc-ASP 155的全长cDNA由955个核苷酸组成，具有多个结构域，包括信号肽（aal-15）、前肽区（aa 16 -45）和典型的催化天冬氨酸结构域（aa 71 -230）。推定的230个氨基酸残基的计算分子量为23.812 Da，理论pI为5.01。Sc-ASP 155 blastp分析显示与寄生和自由生活线虫的天冬氨酸蛋白酶具有40-62%的氨基酸序列同一性。表达分析表明，sc-asp 155基因在线虫寄生初期表达上调，尤其是在13条消化道和6 h诱导期。序列比较表明，Sc-ASP 155属于天冬氨酸蛋白酶家族，系统发育分析表明，Sc-ASP 155与Sc-ASP 113聚在一起。原位杂交显示sc-asp 155在亚腹侧细胞中表达。此外，我们确定sc-asp 155在S.果荚类同源模建表明，Sc-ASP 155具有典型的天冬氨酸蛋白酶结构。Sc-ASP 155的表达上调表明该蛋白酶可能在寄生过程中发挥作用。本研究中，我们克隆了该基因，并测定了胃蛋白酶样天冬氨酸蛋白酶Sc-ASP 155在S.果荚类(C)2012爱思唯尔有限公司版权所有。
Steinernema carpocapsae is an insect parasitic nematode associated with the bacterium Xenorhabdus nematophila. These symbiotic complexes are virulent against the insect host. Many protease genes were shown previously to be induced during parasitism, including one predicted to encode an aspartic protease, which was cloned and analyzed in this study. A cDNA encoding Sc-ASP155 was cloned based on the EST fragment. The full-length cDNA of Sc-ASP155 consists of 955 nucleotides with multiple domains, including a signal peptide (aal-15), a pro-peptide region (aa16-45), and a typical catalytic aspartic domain (aa71-230). The putative 230 amino acid residues have a calculated molecular mass of 23.812 Da and a theoretical pl of 5.01. Sc-ASP155 blastp analysis showed 40-62% amino acid sequence identity to aspartic proteases from parasitic and free-living nematodes. Expression analysis showed that the sc-asp155 gene was up-regulated during the initial parasitic stage, especially in 13 gut and 6 h induced nematodes. Sequence comparison revealed that Sc-ASP155 was a member of an aspartic protease family and phylogenetic analysis indicated that Sc-ASP155 was clustered with Sc-ASP113. In situ hybridization showed that sc-asp155 was expressed in subventral cells. Additionally, we determined that sc-asp155 is a single-copy gene in S. carpocapsae. Homology modeling showed that Sc-ASP155 adopts a typical aspartic protease structure. The up-regulated Sc-ASP155 expression revealed that this protease could play a role in the parasitic process. In this study, we have cloned the gene and determined the expression of the pepsin-like aspartic protease Sc-ASP155 in S. carpocapsae. (C) 2012 Elsevier B.V. All rights reserved.