[1]颜欣欣,杭建雄,刘云秋,等.锡兰钩虫TIMP基因的序列分析与原核表达[J].中国预防兽医学报,2019,(09):969-972.[doi:0.3969/j.issn.1008-0589.201901004]
 YAN Xin-xin,HANG Jian-xiong,LIU Yun-qiu,et al.Sequence analysis and prokaryotic expression of Ancylostoma ceylanicum TIMP[J].Chinese journal of preventive veterinary medicine,2019,(09):969-972.[doi:0.3969/j.issn.1008-0589.201901004]
点击复制

锡兰钩虫TIMP基因的序列分析与原核表达()
分享到:

《中国预防兽医学报》[ISSN:1008-0589/CN:23-1417/S]

卷:
期数:
2019年09
页码:
969-972
栏目:
研究简报
出版日期:
2019-10-25

文章信息/Info

Title:
Sequence analysis and prokaryotic expression of Ancylostoma ceylanicum TIMP
文章编号:
1008-0589(2019)09-0969-04
作者:
 

颜欣欣杭建雄刘云秋黄 岳冉荣坤赵 琪何 龙李 秀刘菊玫李国清*

 (华南农业大学 兽医学院,广东 广州 510642)
Author(s):
 

YAN Xin-xin HANG Jian-xiong LIU Yun-qiu HUANG Yue RAN Rong-kun ZHAO QiHE Long LI xiu LIU Ju-mei LI Guo-qing*

 (College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China)
关键词:
锡兰钩虫金属蛋白酶组织抑制剂基因克隆原核表达序列分析
Keywords:
Ancylostoma ceylanicum  tissue inhibitor of metalloproteinase (TIMP)  cloning  prokaryotic expression phylogenetic analysis
分类号:
S852.7
DOI:
0.3969/j.issn.1008-0589.201901004
文献标志码:
A
摘要:
为表达锡兰钩虫金属蛋白酶组织抑制剂(Ace-TIMP)重组蛋白,本实验根据GenBank中锡兰钩虫TIMP (ANCCEY-04405)基因序列设计引物扩增犬源锡兰钩虫Ace-TIMP基因,利用生物信息学软件对其序列鉴定;根据编码的氨基酸序列进行遗传进化关系分析;构建其重组表达质粒pET-32a-Ace-TIMP-MP,转化大肠杆菌,并经诱导表达。结果显示,Ace-TIMP基因ORF为648 bp,编码215个氨基酸,包含16个氨基酸的信号肽,具有TIMP家族Cys-X-Cys特征性序列。进化树分析显示Ace-TIMP与犬钩虫TMP-2位于同一分支。经诱导表达与纯化,获得了分子质量约为42 ku的可溶性融合蛋白。本实验克隆了Ace-TIMP基因并利用原核表达系统表达了可溶性的重组蛋白,为Ace-TIMP生物学活性的研究和免疫潜能的评价奠定了基础。
  
Abstract:
To prepare recombinant Ancylostoma ceylanicum tissue inhibitor of metalloproteinase (Ace-TIMP), primers were designed according to the sequence in GenBank (ANNECY-04405) to amplify the Ace-TIMP gene from dog-derived parasites. Bioinformatic softwares were used to identify the amplified sequence and phylogenetic analysis was conducted based on the deduced amino acid sequences. A plasmid for expressing Ace-TIMP (pET-32a-Ace-TIMP-MP) in E.coli was constructed to express recombinant protein. Results showed that the ORF of Ace-TIMP is 648bp and encodes 215 amino acids. Ace-TIMP contains a signal peptide with 16 amino acids and the characteristic sequence of TIMP family (Cys-X-Cys). Ace-TIMP is in the same branch with A.caninum TMP-2 in the phylogenetic tree. Finally, soluble fusion protein with molecular weight of about 42ku was expressed and purified from the induced E.coli. In this study, the coding sequence of Ace-TIMP was cloned and expressed through a prokaryotic expression system. The data laid a foundation for further study on biological activities of the protein and evaluation of its immunogenic potential.

参考文献/References:

[1] Thompson R C. Neglected zoonotic helminths: Hymenolepis nana, Echinococcus canadensis and Ancylostoma ceylanicum [J]. Clin Microbiol Infect, 2015, 21(5): 426-432.
[2]Traub R J. Ancylostoma ceylanicum, a re-emerging but neglected parasitic zoonosis [J]. Int J Parasitol, 2013, 43(12-13): 1009-1015.
[3]Yoshikawa M, Ouji Y, Hirai N, et al. Ancylostoma ceylanicum, novel etiological agent for traveler’s diarrhea-eport of four Japanese patients who returned from Southeast Asia and Papua New Guinea [J]. Trop Med Health, 2018, 46: 6.
[4]Bradbury R S, Hii S F, Harrington H, et al. Ancylostoma ceylanicum hookworm in the Solomon Islands [J]. Emerg Infect Dis, 2017, 23(2): 252-257.
[5]Smout F A, Skerratt L F, Johoson C N, et al. The hookworm Ancylostoma ceylanicum: An emerging public health risk in Australian tropical rainforests and Indigenous communities [J]. One Health, 2017, 3: 66-69.
[6]Keiser J, Utzinger J. Efficacy of current drugs against soil-transmitted helminth infections: systematic review and meta-analysis [J]. JAMA, 2008, 299: 1937-1948.
[7]Arpino V, Brock M, Gill S E. The role of TIMPs in regulation of extracellular matrix proteolysis [J]. Matrix Biol, 2015, 44-46: 247-254.
[8]Brew K, Nagase H. The tissue inhibitors of metalloproteinases (TIMPs): an ancient family with structural and functional diversity [J]. Biochim Biophys Acta, 2010, 1803(1): 55-71.
[9]Wei Shuo, Xie Zhi-hong, Filenova E, et al. Drosophila TIMP is a potent inhibitor of MMPs and TACE: similarities in structure and function to TIMP-3 [J]. Biochem, 2003, 42(42): 12200- 12207.
[10]Zhan Bin, Badamchian M, Bo Mei-hua, et al. Molecular cloning and purification of Ac-TMP, a developmentally regulated putative tissue inhibitor of metalloprotease released in relative abundance by adult Ancylostoma hookworms [J]. Am J Trop Med Hyg, 2002, 66(3): 238-244.
[11]Cuellar C, Wu Wen-hui, Mendez S, et al. The hookworm tissue inhibitor of metalloproteases (Ac-TMP-1) modifies dendritic cell function and induces generation of CD4 and CD8 suppressor T cells [J]. PLoS Negl Trop Dis, 2009, 3(5): e439.
[12]Zhan Bin, Gupta R, Wong S P, et al. Molecular cloning and characterization of Ac-TMP-2, a tissue inhibitor of metalloproteinase secreted by adult Ancylostoma caninum [J]. Mol Biochem Parasitol, 2008, 162(2): 142-148.
[13]Mitreva M, McCarter J P, Arasu P, et al. Investigating hookworm genomes by comparative analysis of two Ancylostoma species [J]. BMC Genomics, 2005, 6: 58.
(本文编辑:李 爽;英文编辑:贾洪林)

相似文献/References:

[1]刘远佳,郑国超,刘 田,等.猫锡兰钩虫rDNA ITS1的扩增及分子鉴定[J].中国预防兽医学报,2013,(10):821.[doi:10.3969/j.issn.1008-0589.2013.10.11]
 LIU Yuan-jia,ZHENG Guo-chao,LIU Tian,et al.Isolation and identification of feline hookworm basedon ribosomal ITS1 sequence analysis[J].Chinese journal of preventive veterinary medicine,2013,(09):821.[doi:10.3969/j.issn.1008-0589.2013.10.11]

更新日期/Last Update: 2019-10-29