Enzyme‐Catalyzed Substrate Attachment to Phage Surfaces for the Selection of Catalytic Activities

Phagemid Vectors for Phage Display: Properties, Characteristics and Construction

“…100 Sunbul et al used the similar strategy to achieve dual display in progeny phages, but by modification of protein pIII of the helper phage. 101 Gao et al constructed vector pCGMT-1b in which pIX and pVII were used as anchor proteins for dual display. This strategy does not affect the infectivity of progeny phages.…”

Bacteriophage Capsid Modification by Genetic and Chemical Methods

“…For example, cyclic peptide modified capsid libraries can be generated by intentionally incorporating cysteine residues to facilitate disulfide bond formation in all strains. , Bicyclic peptide libraries can be generated by genetically incorporating three cystines into the capsid gene following reaction with a reagent containing three thiol-reactive groups after phage capsid assembly. − Forming peptides with nonreducible bonds has been achieved in libraries by utilizing enzymes to catalyze peptide cross-linking or noncanonical amino acids that can undergo specific covalent reactions. , In vitro biopanning assays have been used to identify modified phages with affinity to a variety of biological and inorganic substances including single-crystal semiconductors, silica, streptavidin, cellulose, and cell surface receptors. , In vivo biopanning assays have been developed to identify modified phages that hone specific tissues in mouse − and human models. Co-display systems of enzymes and substrates have been developed to identity catalytically active modified capsids during biopanning. − …”

Expanding the Chemical Diversity of Genetically Encoded Libraries

“…Although there are relatively few reports of enzymatic diversification of GELs, enzymes are attractive tools for mild, biocompatible chemistry that are gaining traction. In early work, the kinases Blk, Lyn, c-Src, and Syk were used to phosphorylate tyrosine residues in a phage display library to enrich for and identify peptide sequences similar to the recognition motifs for these kinases. , Since then, several other reports have also used GELs to enrich for substrate sequences for kinases, proteases, and other enzymes. ,− Alternatively, in an approach to create more metabolically stable peptides, several libraries have been “pre-selected” against proteases before target panning to enrich for protease resistant and thus serum stable peptides. , Additionally, sortase A from Staphylococcus aureus has been used to diversify phage particles with synthetic peptides (Figure C.2); phosphorylated and biotinylated peptides, as well as prefolded GFP, all containing LPXTG recognition motifs, were site-specifically ligated onto N-terminal glycines in the M13 phage pIII coat protein. In the same work, another Sortase variant from Streptococcus pyogenes was also used to modify N-terminal alanines of high copy pVIII coat proteins with a fluorophore and single domain antibody .…”