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重组鸡β-防御素2蛋白的表达与生物学活性鉴定 被引量：2

Recombination expression and biological analysis of chicken β-defensin 2
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摘要 为探讨鸡β-防御素2(Av BD2)的生物学特性,研究将Av BD2基因克隆到原核表达载体p Pro EX HTa的Eco RⅠ和XhoⅠ双酶切位点上,构建重组表达质粒p Pro EX-Av BD2,将重组质粒转化大肠杆菌Rosetta感受态细胞,用IPTG对其菌液进行诱导表达。Tricine-SDS-PAGE电泳结果表明,His-Av BD2重组蛋白分子质量约12 ku,与预期一致。将该重组蛋白纯化后,通过菌落计数法测定其体外抗肠炎沙门氏菌、四联球菌、大肠杆菌和枯草芽孢杆菌等抑菌活性,盐离子浓度对其抗菌活性的影响及其溶血活性。结果显示,Av BD2重组蛋白对所测定的4株细菌有显著抗菌活性,高盐浓度显著降低其抗菌活性。该重组蛋白溶血活性极低。 The objective of the study was to investigate bioactivity of chicken avian β-defensin （AvBD） 2 protein. The cDNA of AvBD2 was cloned into EcoR I and Xho I sites of pProEX HTa vector to construct recombinant plasmid pProEX-AvBD2. The recombinant piasmid was transformed into E. coli Rosetta and the bacteria was induced with IPTG. It was demonstrated by Tricine-SDS-PAGE that a 12 kD protein was highly expressed. Antimicrobial activity of the recombinant fusion protein against Salmonella enteritidis, Bacillus subtilis, Micrococcus tetragenus, and E. coll. was measured in vitro. In addition, effect of ionic strength on the antibacterial activity, and hemolytic activity of the recombinant fusion protein were investigated. The recombinant protein exhibited significant antimicrobial activity, the antibacterial activity of AvBD2 significantly decreased in high salt concentration. In addition, recombinant chicken AvBD2 prorein showed little hemolytic activity against chicken erythrocytes.
作者 马得莹 张婷婷 徐杨 高梦莹 王秋玲 徐倩倩 韩宗玺 刘胜旺
机构地区 东北农业大学动物科学技术学院 中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室禽传染病研究室
出处 《东北农业大学学报》 CAS CSCD 北大核心 2015年第8期56-60,108,共6页 Journal of Northeast Agricultural University
基金 教育部博士点基金项目(20122325110016) 哈尔滨市科技创新人才研究专项资金项目(2013RFXXJ019) 黑龙江省应用技术研究与开发计划项目(PC13S02)
关键词 鸡 AvBD2 重组蛋白 抗菌活性 chicken β-defensin 2 recombinant protein antimicrobial activity
分类号 S831 [农业科学—畜牧学] Q786 [生物学—分子生物学]
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参考文献19

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二级参考文献28

  • 1周财源 张名岳 马得莹 等.鹅β-防御素10基因的分离、鉴定与组织分布分析[J].中国农业科学,2011,:51-58.
  • 2Ma DY, Wang RQ, Liao WY, et al. Identification and characterization of a novel Antibacterial peptide, avian B-defensin 2 from ducks. J Microbiol, 2009, 47(5): 610-618.
  • 3Ma DY, Liao WY, Wang RQ, et al. Two novel duck antibacterial peptides, avian 13-defensins 9 and 10, with antimicrobial activity. J Microbiol Biotechnol, 2009, 19(11): 1447-1455.
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  • 5Sugiarto H, Yu PL. Avian antimicrobial peptides: the defense role of beta-defensins. Biochem Biophys Res Commun, 2004, 323(3): 721-727.
  • 6Sugiarto H, Yu PL. Identification of three novel ostricacins: an update on the phylogenetic perspective of beta-defensins. Int J Antimicrob Agents, 2006, 27(3): 229-235.
  • 7Thouzeau C, Maho YL, Froget G, et al. Avain 13-defensins in preserved stomach contents of the king penguin, Aptenodytes patagonicus. J Biol Chem, 2003, 278(51): 51053-51058.
  • 8Milona P, Townes CL, Bevan RM, et al. The chicken host peptides, gallinacins 4, 7, and 9 have antimicrobial activity against Salmonella serovars. Biochem Biophys Res Commun, 2007, 356(1): 1 ~O--17,4.
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  • 10Ma DY, Liu SW, Han ZX, et al. Expression and characterization of recombinant gallinacin-9 and gallinacin-8 in Escherichia coll. Protein Expr Purif, 2008, 58(2): 284-291.

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东北农业大学学报
2015年 第8期

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