Abstract
BackgroundPhage display is a platform for selection of specific binding molecules and this is a clear-cut motivation for increasing its performance. Polypeptides are normally displayed as fusions to the major coat protein VIII (pVIII), or the minor coat protein III (pIII). Display on other coat proteins such as pVII allows for display of heterologous peptide sequences on the virions in addition to those displayed on pIII and pVIII. In addition, pVII display is an alternative to pIII or pVIII display.Methodology/Principal FindingsHere we demonstrate how standard pIII or pVIII display phagemids are complemented with a helper phage which supports production of virions that are tagged with octa FLAG, HIS6 or AviTag on pVII. The periplasmic signal sequence required for pIII and pVIII display, and which has been added to pVII in earlier studies, is omitted altogether.Conclusions/SignificanceTagging on pVII is an important and very useful add-on feature to standard pIII and pVII display. Any phagemid bearing a protein of interest on either pIII or pVIII can be tagged with any of the tags depending simply on choice of helper phage. We show in this paper how such tags may be utilized for immobilization and separation as well as purification and detection of monoclonal and polyclonal phage populations.
Highlights
Phage display is a platform for selection of binders with affinity for specific target molecules, and exhibits high versatility with respect to target discovery [1]
With the exception of pVI display, which has been evaluated for use with cDNA libraries [5], it is common to the majority of phage display protocols that the heterologous peptide is placed in-frame between an N-terminal signal sequence and the mature form of the viral capsid protein
The AviTag-virions were absorbed to streptavidin (SA) coated beads, and bound virions detected by an anti-M13-HRP monoclonal antibody
Summary
Phage display is a platform for selection of binders with affinity for specific target molecules, and exhibits high versatility with respect to target discovery [1]. In both cases, libraries of polypeptides are created as fusions to phage coat proteins that are solvent exposed [2,3].The wt filamentous phage virions M13, fd and f1 have about 2,700 copies of the major coat protein pVIII, and in addition, express approximately 3–5 copies each of pIII, pVI, pVII and pIX; pIII and pVI on one virion tip and pVII and pIX on the other [4]. PVII display is an alternative to pIII or pVIII display
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