Role of Capsid Structure and Membrane Protein Processing in Determining the Size and Copy Number of Peptides Displayed on the Major Coat Protein of Filamentous Bacteriophage.

Filamentous bacteriophage virions can be engineered to display small
foreign peptides in the N-terminal regions of all 2700 copies of the major
coat protein (pVIII), but larger peptides can be accommodated only in
hybrid virions, in which modified and wild-type coat protein subunits are
interspersed. The copy number of peptides accepted in hybrid virions is
generally believed to be related to peptide size: the larger the insert, the
lower the number of modified coat protein subunits in the assembled
virion. However, we show here that some large peptides can be displayed
at a much higher copy number than smaller ones and that some relatively
small peptides are poorly displayed, if at all, in hybrid virions. X-ray
diffraction studies of a recombinant virion together with model building
experiments with peptide and protein epitopes of known structure
demonstrated that it is feasible to accommodate much larger structures,
without perturbation of the capsid protein packing, than it has proved
possible to generate in vivo. We show further that the insertion of certain
peptides greatly slowed or even prevented the processing of the pVIII
pro-coat by leader peptidase at the inner membrane of the Escherichia coli
cell. A good correlation was found between the effect of the insert on the
rate of the processing of the pro-coat, an essential step in virus assembly,
and the number of the mature but modified proteins in the subsequently
assembled hybrid virion. These results have important implications for the
design of peptide display systems based on filamentous bacteriophage.