Selection of novel ligands from a whole-molecule randomly mutated C5a library

S. A. Cain, D. M. Williams, V. Harris, P. N. Monk

    Research output: Contribution to journalArticlepeer-review


    Novel antagonists of the proinflammatory leukocyte chemoattractant C5a have been produced from a phage display library of whole-molecule random mutants. The cDNA for the inflammatory polypeptide C5adR74 was used as template in a PCR reaction doped with the mutagenic nucleoside triphosphates dPTP {dP: 6-(2-deoxy-β-D-ribofu-ranosyl)-3,4-dihydro-8H-pyrimido-[4,5-c][1,2] oxazin-7-one} and 8-oxodGTP (8-oxodG: 8-oxo-2′-deoxyguanosine) to allow the introduction of mutations in a highly controlled manner throughout the cDNA. The resultant library of mutants was displayed on bacteriophage M13 using a jun/fos linker sequence. Functional polypeptides were isolated by several rounds of selection against the receptor for C5a expressed on the surface of CHO cells. From this selection procedure, a limited number of variants of C5adR74 were obtained. When expressed as free polypeptide, the binding affinities of the selected C5adR74 sequences were increased 5-fold relative to wild-type protein. Site-directed mutagenesis of the C-terminus of these variants resulted in the production of antagonists of C5adR74 activity.
    Original languageEnglish
    Pages (from-to)189-193
    Number of pages4
    JournalProtein engineering
    Issue number3
    Publication statusPublished - 2001


    • C5a
    • Phage display
    • Polymerase chain reaction
    • Pyrimidine analogue
    • Random mutagenesis


    Dive into the research topics of 'Selection of novel ligands from a whole-molecule randomly mutated C5a library'. Together they form a unique fingerprint.

    Cite this