Mutagenic Analysis and Localization of a Highly Conserved Epitope Near the Amino-Terminal End of the Citrus Tristeza Closterovirus Capsid Protein
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AuthorPappu, H. R.; Pappu, S. S.; Kano, T.; Koizumi, M.; Cambra, M.; Moreno, P.; Su, H. J.; Garnsey, S. M.; Lee, R. F.; Niblett, C. L.
Cita bibliográficaPappu, H. R., Pappu, S. S., Kano, T., Koizumi, M., Cambra, M., Moreno, P., Su, H. J., Garnsey, S. M., Lee, R. F., Niblett, C. L. (1995). Mutagenic Analysis and Localization of a Highly Conserved Epitope Near the Amino-Terminal End of the Citrus Tristeza Closterovirus Capsid Protein. Phytopathology, 85(10), 1311-1315.
The monoclonal antibody (MAb) 3DF1 is the first commercially available citrus tristeza closterovirus (CTV)-specific MAb. It detects a broad spectrum of CTV isolates from various parts of the world. To precisely map the antigenic determinant recognized by 3DF1, the capsid protein (CP) genes of four 3DF1-nonreactive isolates were cloned as complementary DNA and their nucleotide sequences determined. Comparison of the deduced CP sequences of the four nonreactive isolates with those of previously sequenced 3DF1-reactive isolates revealed differences at three positions near their amino terminal ends. The amino acids Asp-2, Lys-13, and Phe-28 were conserved in all the 3DF1-reactive isolates, but they were replaced by Gly, Thr/Asp, and Tyr, respectively, in the CPs of the nonreactive isolates. Site-specific mutations were introduced into the cloned CP genes of the 3DF1-nonreactive isolate B215 and the 3DF1-reactive isolate T36. The serological reactivities of the wild-type and mutant CPs of B215 and T36 expressed as recombinant fusion proteins in Escherichia coli were evaluated by Western blot analysis. A point mutation (A-->G) resulting in an Asp-->Gly change at amino acid position 2 of the CP of isolate T36 abolished the reactivity with the MAb, whereas a reverse mutation resulting in a Gly-->Asp change at the same position conferred reactivity on the CP of the nonreactive B215 isolate. The implications of the observed antigenic diversity on virus detection are discussed.