Comparison of the Nucleic Acid- and NTP-Binding Properties of the Movement Protein of Cucumber Mosaic Cucumovirus and Tobacco Mosaic Tobamovirus

Abstract

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 intoEscherichia coli,isolating theE. 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 0.4MNaCl, respectively. Both MPs could bind GTP strongly and UTP weakly, but not ATP or CTP. The CMV 3a protein expressed either inE. coliorin plantafrom 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 Ni2+-nitrilotriacetate. The soluble, His-tagged 3a proteins, affinity-purified fromE. coliand zucchini squash, both were able bind CMV RNA 3in vitro.