title: Identification and characterization of mutant clones with enhanced propagation rates from phage-displayed peptide libraries.
abstract: A target-unrelated peptide (TUP) can arise in phage display selection experiments as a result of a propagation advantage exhibited by the phage clone displaying the peptide. We previously characterized HAIYPRH, from the M13-based Ph.D.-7 phage display library, as a propagation-related TUP resulting from a G→A mutation in the Shine-Dalgarno sequence of gene II. This mutant was shown to propagate in Escherichia coli at a dramatically faster rate than phage bearing the wild-type Shine-Dalgarno sequence. We now report 27 additional fast-propagating clones displaying 24 different peptides and carrying 14 unique mutations. Most of these mutations are found either in or upstream of the gene II Shine-Dalgarno sequence, but still within the mRNA transcript of gene II. All 27 clones propagate at significantly higher rates than normal library phage, most within experimental error of wild-type M13 propagation, suggesting that mutations arise to compensate for the reduced virulence caused by the insertion of a lacZα cassette proximal to the replication origin of the phage used to construct the library. We also describe an efficient and convenient assay to diagnose propagation-related TUPS among peptide sequences selected by phage display.
authors: Department of Chemistry, Stonehill College, 320 Washington Street, Easton, MA 02357, USA.
authors: Kieu T H
authors: KT
authors: Nguyen
authors: Department of Chemistry, Stonehill College, 320 Washington Street, Easton, MA 02357, USA.
authors: Marta A
authors: MA
authors: Adamkiewicz
authors: Department of Chemistry, Stonehill College, 320 Washington Street, Easton, MA 02357, USA.
authors: Lauren E
authors: LE
authors: Hebert
authors: Department of Chemistry, Stonehill College, 320 Washington Street, Easton, MA 02357, USA.
authors: Emily M
authors: EM
authors: Zygiel
authors: Department of Chemistry, Stonehill College, 320 Washington Street, Easton, MA 02357, USA.
authors: Holly R
authors: HR
authors: Boyle
authors: Department of Chemistry, Stonehill College, 320 Washington Street, Easton, MA 02357, USA.
authors: Christina M
authors: CM
authors: Martone
authors: Department of Chemistry, Stonehill College, 320 Washington Street, Easton, MA 02357, USA.
authors: Carola B
authors: CB
authors: Meléndez-Ríos
authors: New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938, USA.
authors: Karen A
authors: KA
authors: Noren
authors: New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938, USA.
authors: Christopher J
authors: CJ
authors: Noren
authors: Department of Chemistry, Stonehill College, 320 Washington Street, Easton, MA 02357, USA. Electronic address: mhall@stonehill.edu.
authors: Marilena Fitzsimons
authors: MF
authors: Hall
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D009154:Mutation; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry / Q000235:genetics*
chemical_list: D019151:Peptide Library; D010455:Peptides
keywords: 5′-Untranslated region; M13; Phage display; Propagation-related TUPs; Target-unrelated peptides