[Cloning, expression and characterization analysis of the arginine deiminase of Streptococcus suis of China isolates].
[Cloning, expression and characterization analysis of the arginine deiminase of Streptococcus suis of China isolates].
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发表时间：
2007-10
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通讯作者：
Jin-qiu Zhang;Chengping Lu
中科院分区：
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作者：
Jin-qiu Zhang;Chengping Lu
PCR分析表明，在所有29个S中存在ad基因。suis菌株，但7株S.马链球菌动物寄生虫菌株。将强毒株SS 2-HA 9801的ad基因片段经限制性内切酶酶切后克隆到pBAD/Myc-HisC载体上，转化宿主菌TOP 10。重组蛋白经L-ararose诱导表达后，经Ni-次氮基三乙酸亲和层析柱纯化，表达量为47000 Da。Western blotting结果表明，该重组蛋白能与抗SS 2-HA 9801全细胞蛋白的多克隆抗体发生反应，表明该重组蛋白具有一定的免疫原性。酶法测定表明，其活性的最适温度为37 ℃，pH为6.5。类特异性抑制剂的研究支持巯基酶与一些金属类特征的分配。
PCR analysis demonstrated the presence of the ad gene in all 29 S. suis strains tested, but none of the seven S. equi subsp. zooepidemicus strains. The fragment of ad gene of virulent isolate SS2-HA9801 was later cloned into pBAD/Myc-HisC vector via restriction endonuclease and then transformed into host strain TOP10. A recombinant protein of 47000Da was highly expressed after induced by L-ararose and purified by Ni-nitrilotriacetic acid affinity chromatography. Western blotting demonstrated that the recombinant protein can reacted to the polyclonal antibody raised against whole-cell protein of SS2-HA9801, which suggested that it possessed some immunogenicity and may be important for further research. Enzymatic assay revealed that the optimum temperature for its activity is 37 degrees C and pH is 6.5. Studies with class-specific inhibitors supported the assignment of a sulfhydryl enzyme with some metallo class characteristics.