Comparison of the nucleic acid- and NTP-binding properties of the movement protein of cucumber mosaic cucumovirus and tobacco mosaic tobamovirus.
The cucumber mosaic virus (CMV) 3a movement protein (MP) was compared directly to the well-characterized tobacco mosaic virus (TMV) 30K MP by cloning the genes encoding these proteins into Escherichia coli, isolating the E. coli-expressed MPs, and characterizing them with regard to RNA- and NTP-binding activities. The two MPs were shown to bind single-stranded RNA and DNA cooperatively, but with no sequence specificity. However, discrete lengths of CMV RNA 3 could be protected against RNase digestion by the CMV 3a protein, indicating that the RNA was not uniformly covered by the MP after cooperative binding. The TMV 30K:RNA complex was more stable in NaCl than the CMV 3a:RNA complex; about 50% of the corresponding complexes were stable in 0.6 and 0.4 M NaCl, respectively. Both MPs could bind GTP strongly and UTP weakly, but not ATP or CTP. The CMV 3a protein expressed either in E. coli or in planta from RNA 3 of CMV was tagged at its C-terminus with six histidine residues, which facilitated its purification by affinity chromatography on a matrix containing Ni(2+)-nitrilotriacetate. The soluble, His-tagged 3a proteins, affinity-purified from E. coli and zucchini squash, both were able bind CMV RNA 3 in vitro.
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Department of Plant Pathology, Cornell University, Ithaca, New York 14853, USA.
Palukaitis
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D001483:Base Sequence; D017799:Cucumovirus / Q000235:genetics / Q000378:metabolism*; D004279:DNA, Viral / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004926:Escherichia coli; D006639:Histidine / Q000378:metabolism; D008969:Molecular Sequence Data; D009711:Nucleotides / Q000378:metabolism*; D053830:Plant Viral Movement Proteins; D010944:Plants / Q000821:virology; D011485:Protein Binding; D012367:RNA, Viral / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D012965:Sodium Chloride / Q000494:pharmacology; D014027:Tobacco Mosaic Virus / Q000235:genetics / Q000378:metabolism*; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D009711:Nucleotides; D053830:Plant Viral Movement Proteins; D012367:RNA, Viral; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D014764:Viral Proteins; D012965:Sodium Chloride; D006639:Histidine