Determination of the complete amino acid sequence for the coat protein of brome mosaic virus by time-of-flight mass spectrometry - Evidence for mutations associated with change of propagation host

Time-of-flight mass spectrometry (TOFMS) has been applied to determine the complete coat protein amino acid sequences of a number of distinct brome mo saic virus (BMV) isolates. Ionization was carried out by both electrospray ionization and matrix-assisted laser desorption/ionization (MALDI), After d etermining overall coat protein masses, the proteins were digested with try psin or Lys-C proteinases, and the digestion products were analyzed in a MA LDI QqTOF mass spectrometer. The N terminus of the coat protein was found t o be acetylated in each BMV isolate analyzed. In one isolate (BMV-Valverde) , the amino acid sequence was identical to that predicted from the cDNA seq uence of the "type" isolate, but deviations from the predicted amino acid s equence were observed for all the other isolates analyzed, When isolates we re propagated in different host taxa, modified coat protein sequences were observed in some cases, along with the original sequence. Sequencing by TOF MS may therefore provide a basis for monitoring the effects of host passagi ng on a virus at the molecular level. Such TOFMS-based analyses assess the complete profiles of coat protein sequences actually present in infected ti ssues. They are therefore not subject to the selection biases inherent in d educing such sequences from reverse-transcribed viral RNA and cloning the r esulting cDNA.