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Journal of Virology, September 2001, p. 8538-8546, Vol. 75, No. 18
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.18.8538-8546.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Biochemical Characterization of the Helper Component of Cauliflower Mosaic Virus

Eugenie Hebrard,1 Martin Drucker,1 Denis Leclerc,2,dagger Thomas Hohn,2 Marilyne Uzest,1 Remy Froissart,1 Jean-Marc Strub,3 Sarah Sanglier,3 Alain van Dorsselaer,3 Andre Padilla,4 Gilles Labesse,4 and Stephane Blanc1,*

Station de Recherches de Pathologie Comparée, UMR 5087, INRA-CNRS-Université Montpellier II, 30380 Saint-Christol-les-Alès,1 Laboratoire de Spectrométrie de Masse Bio-Organique, 67087 Strasbourg Cedex 2,3 and Centre de Biochimie Structurale, INSERM U414, CNRS UMR 5048-Université Montpellier I Faculté de Pharmacie, 34060 Montpellier,4 France, and Friedrich Miescher Institut, CH-4002 Basel, Switzerland2

Received 26 March 2001/Accepted 5 June 2001

The helper component of Cauliflower mosaic virus is encoded by viral gene II. This protein (P2) is dispensable for virus replication but required for aphid transmission. The purification of P2 has never been reported, and hence its biochemical properties are largely unknown. We produced the P2 protein via a recombinant baculovirus with a His tag fused at the N terminus. The fusion protein was purified by affinity chromatography in a soluble and biologically active form. Matrix-assisted laser desorption time-of-flight mass spectrometry demonstrated that P2 is not posttranslationally modified. UV circular dichroism revealed the secondary structure of P2 to be 23% alpha -helical. Most alpha -helices are suggested to be located in the C-terminal domain. Using size exclusion chromatography and aphid transmission testing, we established that the active form of P2 assembles as a huge soluble oligomer containing 200 to 300 subunits. We further showed that P2 can also polymerize as long paracrystalline filaments. We mapped P2 domains involved in P2 self-interaction, presumably through coiled-coil structures, one of which is proposed to form a parallel trimer. These regions have previously been reported to also interact with viral P3, another protein involved in aphid transmission. Possible interference between the two types of interaction is discussed with regard to the biological activity of P2.

* Corresponding author. Mailing address: Station de Recherches de Pathologie Comparée, UMR 5087, INRA-CNRS-Université Montpellier II, 30380 Saint-Christol-les-Alès, France. Phone: 33 4 66 78 37 15. Fax: 33 4 66 52 46 99. E-mail: blanc{at}ensam.inra.fr.

dagger Present address: Centre de Recherche en Infectiologie PavillonCHUL, Université Laval, Ste-Foy, P.Q. G1V 4G2, Canada.

Journal of Virology, September 2001, p. 8538-8546, Vol. 75, No. 18
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.18.8538-8546.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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