Identification of regions of brome mosaic virus coat protein chemically cross-linked in situ to viral RNA

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Abstract
RNA-protein cross-links were introduced into brome mosaic virus in situ by using the heterobifunctional agent p-azidophenylglyoxal. An improved RNA isolation method, without phenol extraction, was used to isolate RNA cross-linked with protein. RNA of the covalently linked complex was acid-digested and the oligonucleotides still attached to protein were 5''-end-labelled with 32P. The complexes were digested with trypsin and the tryptic peptides were purified by reversed-phase high-performance liquid chromatography. Amino acid analyses of cross-linked tryptic peptides revealed that out of the total 188 amino acids of brome mosaic virus coat protein only the 80 N-terminal amino acids are involved in the interaction with viral RNA. These results are discussed in connection with predicted secondary of the coat protein. Both .alpha. helix (for amino acids 11-19) and other structures (between amino acids 20 and 80) are implicated in the coat protein-viral RNA interactions.