Single-chain Fv phage display propensity exhibits strong positive correlation with overall expression levels

Abstract

Abstract Background Single chain Fvs (scFvs) are widely applied in research, diagnostics and therapeutic settings. Display and selection from combinatorial libraries is the main route to their discovery and many factors influence the success of this process. They exhibit low thermodynamic stability, resulting in low levels of premature cytosolic folding or aggregation which facilitates sec YEG-mediated translocation and phage in E. coli. However, there is little data analysing how this is related to and influenced by scFv protein expression. Results We characterised the relationship between overall scFv expression and display propensity for a panel of 15 anti-tetanus toxin scFvs and found a strong positive correlation (Rho = 0.88, p Conclusion We conclude that in the absence of premature cytosolic aggregation or folding, the propensity of a scFv to be displayed on phage is directly related to its overall expression level and is thus indirectly influenced by factors such as codon bias, mRNA abundance or putative DNA motifs affecting expression. This suggests that scFvs capable of high overall expression and display levels may not produce high yields of non phage-fused soluble protein in either the periplasmic or extracellular fractions of E. coli. This should be considered when screening clones selected from combinatorial libraries for further study. The nucleotide and amino acid sequences of the anti-tetanus toxin scFvs have been deposited in the EMBL data base: accession numbers-C1: AM749134, C2: AM749135, C3: AM749136, C4: AM749137, C5: AM749138, N1: AM749139, N2: AM749140, N3: AM749141, N4: AM749142, N5: AM749143 J1; AM749144, J2: AM749145, J3: AM749146, J4: AM749147, J5: AM749148.