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J. Biol. Chem., Vol. 282, Issue 6, 3458-3464, February 9, 2007

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Plant Transformation by Agrobacterium tumefaciens

MODULATION OF SINGLE-STRANDED DNA-VirE2 COMPLEX ASSEMBLY BY VirE1^*

Daphna Frenkiel-Krispin, Sharon Grayer Wolf, Shira Albeck^¶, Tamar Unger^¶, Yoav Peleg^¶, Jossef Jacobovitch^¶, Yigal Michael^¶, Shirley Daube, Michal Sharon^||, Carol V. Robinson^||, Dmitri I. Svergun^**¶¶, Deborah Fass, Tzvi Tzfira, and Michael Elbaum¹

From the Departments of Materials and Interfaces, Chemical Research Support, and Structural Biology and the ^¶Israel Structural Proteomics Center, Weizmann Institute of Science, Rehovot 76100, Israel, the ^||Department of Chemistry, Cambridge University, Lensfield Road, Cambridge CB2 1EW, United Kingdom, the ^**European Molecular Biology Laboratory, Hamburg Outstation, Notkestrasse 85, D-22603 Hamburg, Germany, the ^¶¶Institute of Crystallography, Russian Academy of Sciences, Leninsky Prospekt 59, 117333 Moscow, Russia, and the Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Ann Arbor, Michigan 48109

Agrobacterium tumefaciens infects plant cells by the transfer of DNA. A key factor in this process is the bacterial virulence protein VirE2, which associates stoichiometrically with the transported single-stranded (ss) DNA molecule (T-strand). As observed in vitro by transmission electron microscopy, VirE2-ssDNA readily forms an extended helical complex with a structure well suited to the tasks of DNA protection and nuclear import. Here we have elucidated the role of the specific molecular chaperone VirE1 in regulating VireE2-VirE2 and VirE2-ssDNA interactions. VirE2 alone formed functional filamentous aggregates capable of ssDNA binding. In contrast, co-expression with VirE1 yielded monodisperse VirE1–VirE2 complexes. Cooperative binding of VirE2 to ssDNA released VirE1, resulting in a controlled formation mechanism for the helical complex that is further promoted by macromolecular crowding. Based on this in vitro evidence, we suggest that the constrained volume of the VirB channel provides a natural site for the exchange of VirE2 binding from VirE1 to the T-strand.

Received for publication, June 1, 2006 , and in revised form, October 10, 2006.

^* This work was supported by the United States-Israel Binational Agricultural Research and Development Fund (BARD) and by the Gerhard M. J. Schmidt Minerva Center for Supramolecular Architecture. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ To whom correspondence should be addressed: Dept. of Materials and Interfaces, Weizmann Inst. of Science, Rehovot 76100, Israel. Tel.: 972-8-9343537; Fax: 972-8-9344138; E-mail: michael.elbaum{at}weizmann.ac.il.

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