Biological technique to evolve proteins using bacteriophages
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Phage display cycle. 1) fusion proteins for a viral coat protein + the gene to be evolved (typically an antibody fragment) are expressed in bacteriophage. 2) the library of phage are washed over an immobilised target. 3) the remaining high-affinity binders are used to infect bacteria. 4) the genes encoding the high-affinity binders are isolated. 5) those genes may have random mutations introduced and used to perform another round of evolution. The selection and amplification steps can be performed multiple times at greater stringency to isolate higher-affinity binders.
Phage display is a laboratory technique for the study of protein–protein, protein–peptide, and protein–DNA interactions that uses bacteriophages (viruses that infect bacteria) to connect proteins with the genetic information that encodes them.[1] In this technique, a gene encoding a protein of interest is inserted into a phage coat protein gene, causing the phage to "display" the protein on its outside while containing the gene for the protein on its inside, resulting in a connection between genotype and phenotype. The proteins that the phages are displaying can then be screened against other proteins, peptides or DNA sequences, in order to detect interaction between the displayed protein and those of other molecules. In this way, large libraries of proteins can be screened and amplified in a process called in vitro selection, which is analogous to natural selection.
The most common bacteriophages used in phage display are M13 and fd filamentous phage,[2][3] though T4,[4] T7, and λ phage have also been used.
^Smith GP (June 1985). "Filamentous fusion phage: novel expression vectors that display cloned antigens on the virion surface". Science. 228 (4705): 1315–7. Bibcode:1985Sci...228.1315S. doi:10.1126/science.4001944. PMID 4001944.
^Kehoe JW, Kay BK (November 2005). "Filamentous phage display in the new millennium". Chem. Rev. 105 (11): 4056–72. doi:10.1021/cr000261r. PMID 16277371.
^Malys N, Chang DY, Baumann RG, Xie D, Black LW (2002). "A bipartite bacteriophage T4 SOC and HOC randomized peptide display library: detection and analysis of phage T4 terminase (gp17) and late sigma factor (gp55) interaction". J Mol Biol. 319 (2): 289–304. doi:10.1016/S0022-2836(02)00298-X. PMID 12051907.
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