Peptide display on live MS2 phage: restrictions at the RNA genome level

The potential of the RNA phage MS2 to accommodate extra amino acids in its major coat protein has been examined. Accordingly, a pentapeptide was encod ed in the genome as an N-terminal extension. In the MS2 crystal structure, this part of the coat protein forms a loop that extends from the outer surf ace of the icosahedral virion. At the RNA level, the insert forms a large l oop at the top of an existing hairpin. This study shows that it is possible to maintain inserts in the coat protein of live phages, However, not all i nserts were genetically stable. Some suffer deletions, while others underwe nt adaptation by base substitutions. Whether or not an insert is stable app ears to be determined by the choice of the nucleic acid sequence used to en code the extra peptide. This effect was not caused by differential translat ion, because coat-protein synthesis was equal in wild-type and mutants. We conclude that the stability of the insert depends on the structure of the l arge RNA hairpin loop, as demonstrated by the fact that a single substituti on can convert an unstable loop into a stable one.