Isolation of fibroblast growth factor receptor binding sequences using evolved phage display libraries.
A fusion protein construct consisting of the short form of human fibroblast growth factor (FGFR) fused to the heavy chain of mouse IgG1 was used to screen four phage display libraries displaying 8, 13, 38 and 43 amino acids at the amino terminus of the bacteriophage M13 gene III minor coat protein. Phage with specific FGFR binding activity were isolated from the 13, 38 and 43 mer libraries. One of the highest affinity phage clones from the 13mer library was chosen to be further evolved by oligonucleotide saturation mutagenesis. We have isolated evolved sequences that have approximately 8 times the relative binding affinity of the parent sequence. The phage clones have a minimum consensus sequence of CR/SXLLXGAPFXXXXC, where X represents positions tolerant of several amino acids. A synthetic peptide based on this sequence specifically inhibits FGF from binding to its receptor in an in vitro ELISA.
McConnell
S J
SJ
Biological Screening Group, Chugai Biopharmaceuticals, Inc., San Diego, CA 92121, USA. smcconnell@chugaibio.com
Thon
V J
VJ
Spinella
D G
DG
D000595:Amino Acid Sequence; D000818:Animals; D001435:Bacteriophages / Q000235:genetics*; D001483:Base Sequence; D001665:Binding Sites; D019556:COS Cells; D003001:Cloning, Molecular; D004247:DNA; D004797:Enzyme-Linked Immunosorbent Assay; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism; D006801:Humans; D007074:Immunoglobulin G / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D017468:Receptors, Fibroblast Growth Factor / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics
D007074:Immunoglobulin G; D017468:Receptors, Fibroblast Growth Factor; D011993:Recombinant Fusion Proteins; D016222:Fibroblast Growth Factor 2; D004247:DNA