Suppr超能文献

中国对虾(凡纳滨对虾)Kazal型蛋白酶抑制剂的表征、动力学及可能功能

Characterization, kinetics, and possible function of Kazal-type proteinase inhibitors of Chinese white shrimp, Fenneropenaeus chinensis.

作者信息

Wang Zong-Heng, Zhao Xiao-Fan, Wang Jin-Xing

机构信息

School of Life Sciences, Shandong University, Jinan 250100, China.

出版信息

Fish Shellfish Immunol. 2009 Jun;26(6):885-97. doi: 10.1016/j.fsi.2009.03.024. Epub 2009 Apr 18.

Abstract

Serine proteinase inhibitor plays an essential role in arthropods by restraining the activities of endogenic or exogenic serine proteinases. Four Kazal-type serine proteinase inhibitors, Fcspi-1-4, from the hepatopancreas of Chinese white shrimp, Fenneropenaeus chinensis, were cloned and identified. The open reading frames (ORFs) of Fcspis are 1389, 1236, 1080, and 939 base pairs, encode the pre-proteins of 462, 411, 359, and 312 amino acids and form the 9, 8, 7, and 6 typical Kazal domains, respectively. When analyzing the amino acid sequences of the four inhibitors, it was found that they might have been derived from the same transcript, which was subjected to alternative splicing, and none of the Kazal domains were identical within each inhibitor. Multiple alignments showed that the Kazal inhibitors were homologous with a conserved motif of Cx(3)Cx(6)VCGSDGxTYx(3)CxLx(5)Cx(5)ITx(6)GC. The results from RT-PCR indicated that the expression of Fcspis as a whole was upregulated by bacterial challenge, no obvious change was noticed after viral challenge, and Fcspi-1 had a similar expression pattern with that of Fcspis. Recombinant FcSPIs were successfully expressed in bacteria and purified for further study. Recombinant FcSPI-1 was sensitive to DTT and had thermal stability. The inhibitory kinetics assay suggested that rFcSPI-1 was a mixed-type fast tight binding inhibitor with inhibitory activities against subtilisin A at a molar ratio of 1:1, 1:2 against proteinase K, and 2:1 against elastase. It can firmly bound to two Gram-positive and one Gram-negative bacteria but without anti-bacterial ability. In addition, it inhibited the activities of both bacterial-secreted proteinases and natural chymotrypsin of Chinese white shrimp, suggesting that FcSPI-1 may participate in the immune defence response by inhibition of bacterial pathogen proteinases and possibly be involved in the regulation of shrimp proteinase activity.

摘要

丝氨酸蛋白酶抑制剂通过抑制内源性或外源性丝氨酸蛋白酶的活性,在节肢动物中发挥着重要作用。从中国对虾(凡纳滨对虾)的肝胰腺中克隆并鉴定出了四种卡扎尔型丝氨酸蛋白酶抑制剂Fcspi - 1 - 4。Fcspis的开放阅读框(ORF)分别为1389、1236、1080和939个碱基对,编码462、411、359和312个氨基酸的前体蛋白,并分别形成9、8、7和6个典型的卡扎尔结构域。在分析这四种抑制剂的氨基酸序列时发现,它们可能来自同一个经过可变剪接的转录本,并且每种抑制剂内的卡扎尔结构域都不相同。多重比对显示,卡扎尔抑制剂与保守基序Cx(3)Cx(6)VCGSDGxTYx(3)CxLx(5)Cx(5)ITx(6)GC同源。RT - PCR结果表明,细菌刺激后Fcspis的整体表达上调,病毒刺激后未观察到明显变化,且Fcspi - 1与Fcspis具有相似的表达模式。重组FcSPIs在细菌中成功表达并纯化用于进一步研究。重组FcSPI - 1对二硫苏糖醇(DTT)敏感且具有热稳定性。抑制动力学分析表明,rFcSPI - 1是一种混合型快速紧密结合抑制剂,对枯草杆菌蛋白酶A的抑制活性摩尔比为1:1,对蛋白酶K为1:2,对弹性蛋白酶为2:1。它能牢固地结合两种革兰氏阳性菌和一种革兰氏阴性菌,但没有抗菌能力。此外,它抑制了细菌分泌的蛋白酶以及中国对虾天然胰凝乳蛋白酶的活性,这表明FcSPI - 1可能通过抑制细菌病原体蛋白酶参与免疫防御反应,并可能参与对虾蛋白酶活性的调节。

文献AI研究员

20分钟写一篇综述,助力文献阅读效率提升50倍。

立即体验

用中文搜PubMed

大模型驱动的PubMed中文搜索引擎

马上搜索

文档翻译

学术文献翻译模型,支持多种主流文档格式。

立即体验