In vitroselection and evolution of functional proteins by using ribosome display

• DOI: 10.1073/pnas.94.10.4937
• Publication Date: 2002-07-26T14:31:39Z
• Authors: Jozef Hanes; Andreas Plückthun
• Keywords: 0301 basic medicine; Protein Folding; Binding Sites; Base Sequence; Transcription, Genetic; Molecular Sequence Data; Protein Engineering; Biological Evolution; Polymerase Chain Reaction; Recombinant Proteins; 03 medical and health sciences; Protein Biosynthesis; Mutagenesis, Site-Directed; Point Mutation; Amino Acid Sequence; RNA, Messenger; Immunoglobulin Fragments; Ribosomes; DNA Primers; Information Systems

We report here a system with which correctly folded complete protein and its encoding mRNA both remain attached to the ribosome can be enriched for ligand-binding properties of native protein. have selected single-chain fragment (scFv) an antibody 10 8 -fold by five cycles transcription, translation, antigen-affinity selection, PCR. The scFv fragments all mutated in vitro acquiring up four unrelated amino acid exchanges over generations, but they remained fully compatible antigen binding. Libraries proteins now screened made evolve cell-free without any transformation or constraints imposed host cell.