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| Expression and Activity Analysis of Non-Structural Protein 2 （NS2） of Bombyx mori Densovirus Zhenjiang Strain | |
| 作者姓名： | ZHAO Pan TANG Shun-ming LIU Tingx LIU Ting QIN Guang-xing GUO Xi-j ie |
| 作者单位： | [1]College of Biotechnology and Environmental Engineering, Jiangsu University of Science and Technology, Zhenjiang 212003, P.R.China [2]Key Laboratory of Silkworm and Mulberry Genetic Improvement, Ministry of Agriculture of China/Sericultural Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang 212018, P.R.China |
| 基金项目： | This work was supported by the National Basic Research Program of China （2005CB 121004）. |
| 摘 要： | The gene of the non-structure protein 2 （NS2） was cloned by PCR from the genome ofBombyx mori densovirus Zhenjiang strain （BmDNV-Z）, inserted into prokaryotic expression vector pET28a to construct recombinant plasmid pET28a-NS2 and then expressed in bacteria Escherichia coli BL21 （DE3）. The expressed recombinant protein was identified by SDS-PAGE and Western blot analysis. Then, the recombinant protein was purified by Ni-NTA column, renatured and tested for enzyme activities. The purified NS2 protein exhibited a helicase activity unwinding double-stranded DNA substrates into single-strand primers, and higher unwinding activity to polarity substrate. Similarly, the purified NS2 protein possessed an ATPase activity and its enzyme activity was 0.276 μmol gg^-1 h^-1 in this study. The results indicated that the non- structure protein which encoded by the gene of BmDNV-Z NS2 possesses the biological activities of helicase and ATPase, and the helicase prefers to polarity substrates. Based on these results, it is speculated that the gene of BmDNV-Z NS2 plays an important role in the viral DNA replication. |
| 关 键 词： | NS2基因 非结构蛋白 活性分析 浓核病毒 镇江 家蚕 蛋白质纯化 原核表达载体 |
Expression and Activity Analysis of Non-Structural Protein 2 (NS2) of Bombyx mori Densovirus Zhenjiang Strain |
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| ZHAO Pan,TANG Shun-ming,LIU Tingx,LIU Ting,QIN Guang-xing,GUO Xi-j ie.Expression and Activity Analysis of Non-Structural Protein 2 (NS2) of Bombyx mori Densovirus Zhenjiang Strain[J].Agricultural Sciences in China,2010,9(12):1821-1828. | |
| Authors: | ZHAO Pan TANG Shun-ming LIU Ting QIN Guang-xing GUO Xi-jie |
| Institution: | 1. College of Biotechnology and Environmental Engineering,Jiangsu University of Science and Technology,Zhenjiang 212003,P.R.China 2. College of Biotechnology and Environmental Engineering,Jiangsu University of Science and Technology,Zhenjiang 212003,P.R.China;Key Laboratory of Silkworm and Mulberry Genetic Improvement,Ministry of Agriculture of China/Sericultural Research Institute,Chinese Academy of Agricultural Sciences,Zhenjiang 212018,P.R.China 3. Key Laboratory of Silkworm and Mulberry Genetic Improvement,Ministry of Agriculture of China/Sericultural Research Institute,Chinese Academy of Agricultural Sciences,Zhenjiang 212018,P.R.China |
| Abstract: | The gene of the non-structure protein 2 (NS2) was cloned by PCR from the genome of Bombyx mori densovirus Zhenjiang strain (BmDNV-Z), inserted into prokaryotic expression vector pET28a to construct recombinant plasmid pET28a-NS2 and then expressed in bacteria Escherichia coli BL21 (DE3). The expressed recombinant protein was identified by SDS-PAGE and Western blot analysis. Then, the recombinant protein was purified by Ni-NTA column, renatured and tested for enzyme activities. The purified NS2 protein exhibited a helicase activity unwinding double-stranded DNA substrates into single-strand primers, and higher unwinding activity to polarity substrate. Similarly, the purified NS2 protein possessed an ATPase activity and its enzyme activity was 0.276 μtmol μg-1 h-1 in this study. The results indicated that the nonstructure protein which encoded by the gene of BmDNV-Z NS2 possesses the biological activities of helicase and ATPase,and the helicase prefers to polarity substrates. Based on these results, it is speculated that the gene of BmDNV-Z NS2 plays an important role in the viral DNA replication. |
| Keywords: | Bombyx mori densovirus non-structure protein expression activity |
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