Structural properties of carnation mottle virus p7 movement protein and its RNA-binding domain.

Abstract

Plant viral movement proteins (MPs) participate actively in the intra- and intercellular movement of RNA plant viruses to such an extent that MP dysfunction impairs viral infection. However, the molecular mechanism(s) of their interaction with cognate nucleic acids are not well understood, partly due to the lack of structural information. In this work, a protein dissection approach was used to gain information on the structural and RNA-binding properties of this class of proteins, as exemplified by the 61-amino acid residue p7 MP from carnation mottle virus (CarMV). Circular dichroism spectroscopy showed that CarMV p7 is an alpha/beta RNA-binding soluble protein. Using synthetic peptides derived from the p7 sequence, we have identified three distinct putative domains within the protein. EMSA showed that the central region, from residue 17 to 35 (represented by peptide p7(17-35)), is responsible for the RNA binding properties of CarMV p7. This binding peptide populates a nascent alpha-helix in water solution that is further stabilized in the presence of either secondary structure inducers, such as trifluoroethanol and monomeric SDS, or RNA (which also changes its conformation upon binding to the peptide). Thus, the RNA recognition appears to occur via an "adaptive binding" mechanism. Interestingly, the amino acid sequence and structural properties of the RNA-binding domain of p7 seem to be conserved among carmoviruses and some other RNA-binding proteins and peptides. The low conserved N terminus of p7 (peptide p7(1-16)) is unstructured in solution. In contrast, the highly conserved C terminus motif (peptide p7(40-61)) adopts a beta-sheet conformation in aqueous solution. Alanine scanning mutagenesis of the RNA-binding motif showed how selected positive charged amino acids are more relevant than others in the RNA binding process and how hydrophobic amino acid side chains would participate in the stabilization of the protein-RNA complex.

• Authors: M Vilar, V Esteve, V Pallas, J F Marcos, E Perez-Paya
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 276 Issue 21 Pg. 18122-9 (May 25 2001) ISSN: 0021-9258 [Print] United States
• PMID: 11279121 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• P7 protein, Carnation mottle virus
• RNA-Binding Proteins
• Viral Proteins
• RNA

Topics

• Amino Acid Sequence
• Binding Sites
• Carlavirus
• Molecular Sequence Data
• Protein Structure, Secondary
• RNA (metabolism)
• RNA-Binding Proteins (genetics, metabolism)
• Viral Proteins (genetics, metabolism)