title: A novel helper phage for HaloTag-mediated co-display of enzyme and substrate on phage.
abstract: Phage display is an established technique for the molecular evolution of peptides and proteins. For the selection of enzymes based on catalytic activity however, simultaneous coupling of an enzyme and its substrate to the phage surface is required. To facilitate this process of co-display, we developed a new helper phage displaying HaloTag, a modified haloalkane dehalogenase that binds specifically and covalently to functionalized haloalkane ligands. The display of functional HaloTag was demonstrated by capture on streptavidin-coated magnetic beads, after coupling a biotinylated haloalkane ligand, or after on-phage extension of a DNA oligonucleotide primer with a biotinylated nucleotide by phi29 DNA polymerase. We also achieved co-display of HaloTag and phi29 DNA polymerase, thereby opening perspectives for the molecular evolution of this enzyme (and others) towards new substrate specificities.
authors.affiliation: Department of Chemistry, KU Leuven, Celestijnenlaan 200G, B-3001 Heverlee, Belgium. Electronic address: wouter.delespaul@chem.kuleuven.be.
authors.forename: Wouter
authors.initials: W
authors.lastname: Delespaul
authors.affiliation: Department of Chemistry, KU Leuven, Celestijnenlaan 200G, B-3001 Heverlee, Belgium. Electronic address: yves.peeters@chem.kuleuven.be.
authors.forename: Yves
authors.initials: Y
authors.lastname: Peeters
authors.affiliation: Rega Institute for Medicinal Research, KU Leuven, Minderbroedersstraat 10, B-3000 Leuven, Belgium. Electronic address: piet.herdewijn@rega.kuleuven.be.
authors.forename: Piet
authors.initials: P
authors.lastname: Herdewijn
authors.affiliation: Department of Chemistry, KU Leuven, Celestijnenlaan 200G, B-3001 Heverlee, Belgium. Electronic address: johan.robben@chem.kuleuven.be.
authors.forename: Johan
authors.initials: J
authors.lastname: Robben
mesh_terms: D001435:Bacteriophages / Q000201:enzymology / Q000378:metabolism*; D001483:Base Sequence; D017931:DNA Primers; D006867:Hydrolases / Q000235:genetics* / Q000378:metabolism; D008024:Ligands; D016296:Mutagenesis; D016133:Polymerase Chain Reaction; D013379:Substrate Specificity
chemical_list: D017931:DNA Primers; D008024:Ligands; D006867:Hydrolases; C046137:haloalkane dehalogenase
keywords: Co-display; HaloTag; Molecular evolution; Phage display; Polymerase; Self-tagging