Molecular characterization of the Rep protein of the blackgram isolate of Indian mungbean yellow mosaic virus

Abstract

The complete nucleotide sequence of the blackgram isolate of mungbean yellow mosaic virus, IMYMV-Bg, which infects legumes in India, was determined and compared at the amino acid level with those of other whitefly-transmitted geminiviruses. The genome organization of IMYMV-Bg was similar to that of the begomoviruses. A unique feature of the genome organization was the sequence divergence of the common region (CR) between DNA-A and DNA-B. In order to understand the mechanism of viral DNA replication, the replication initiator protein, Rep, of IMYMV-Bg was overexpressed in E. coli. The recombinant and refolded Rep bound to CR-sequences of IMYMV-Bg in a specific manner. In this study, evidence is presented for ATP-upregulated cleavage function and ATP-mediated conformational change of Rep. It is hypothesized that, although ATP is not required for cleavage, ATP-mediated conformational changes may result in better access of Rep to the DNA-cleavage site. Evidence is also presented for a site-specific topoisomerase function of Rep, which has not been demonstrated before. The Rep protein can be classified as a type-I topoisomerase because of its nicking activity and sensitivity towards camptothecin, a topoisomerase type-I inhibitor.