NDLI: Isolation, purification, and identification of the virulence protein VirE2 from Agrobacterium tumefaciens

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Isolation, purification, and identification of the virulence protein VirE2 from Agrobacterium tumefaciens

Content Provider	Springer Nature Link
Author	Volokhina, I. V. Sazova, I. A. Velikov, V. A. Chumakov, M. I.
Copyright Year	2005
Abstract	Bacteria of the genus Agrobacterium are capable of transferring a fragment of their Ti-plasmid T-DNA, in a complex with the proteins VirE2 and VirD2, into the nuclei of plant cells and incorporating it into the chromosome of the host. The mechanisms of T-DNA transportation through the membrane and cytoplasm of the plant cell are unknown. The aim of this work was isolation of the virulence protein VirE2 for studying its role in T-DNA transportation through the membrane and cytoplasm of eukaryotic cells. For VirE2 accumulation, the virE2 gene was cloned into plasmid pQE31. VirE2 was isolated from the cells of E. coli strain XL1-blue, containing the recombinant plasmid pQE31-virE2. The cells were disrupted ultrasonically, and the protein, with six histidine residues at the N-end, was isolated by means of affinity chromatography on a Ni-NTA-superose column. The purified protein was tested by the immunodot method using polyclonal rabbit antibodies and anti-VirE2 miniantibodies. The ability of the recombinant protein VirE2 to bind to single-stranded DNA was judged from the formation of complexes detected by electrophoresis in agarose gel. Thus, we isolated, purified, and partially characterized the Agrobacterium tumefaciens virulence protein VirE2, which is capable of binding to single-stranded T-DNA upon transfer to the plant cell.
Starting Page	80
Ending Page	85
Page Count	6
File Format	PDF
ISSN	00262617
Journal	Microbiology
Volume Number	74
Issue Number	1
e-ISSN	16083237
Language	English
Publisher	Nauka/Interperiodica
Publisher Date	2005-01-01
Publisher Place	Moscow
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	Agrobacterium tumefaciens virulence protein VirE2 cloning in E. coli affinity chromatography phage display miniantibodies Medical Microbiology Microbiology
Content Type	Text
Resource Type	Article
Subject	Microbiology Applied Microbiology and Biotechnology

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