A1 Refereed original research article in a scientific journal
Synonymous Codons and Hydrophobicity Optimization of Post-translational Signal Peptide PelB Increase Phage Display Efficiency of DARPins
Authors: Kulmala Antti, Lappalainen Matias, Lamminmäki Urpo, Huovinen Tuomas
Publisher: AMER CHEMICAL SOC
Publication year: 2022
Journal: ACS Synthetic Biology
Journal name in source: ACS SYNTHETIC BIOLOGY
Journal acronym: ACS SYNTH BIOL
Number of pages: 8
ISSN: 2161-5063
eISSN: 2161-5063
DOI: https://doi.org/10.1021/acssynbio.2c00260
Web address : https://pubs.acs.org/doi/10.1021/acssynbio.2c00260
Self-archived copy’s web address: https://research.utu.fi/converis/portal/detail/Publication/176747174
DsbA leader peptide targets proteins for cotransla-tional translocation by signal recognition particle (SRP) pathway and has been the standard signal sequence for filamentous phage display of fast-folding Designed Ankyrin Repeat Proteins (DARPins). In contrast, translocation of DARPins via the post-translational pathway, for example, with the commonly used PelB leader, has been reported to be highly inefficient. In this study, two PelB signal sequence libraries were screened covering different regions of the leader peptide for identifying mutants with improved display of DARPins on phage. A PelB variant with the most favorable combination of synonymous mutations in the n-region and hydrophobic substitutions in the h-region increased the display efficiency of a DARPin library 44-and 12-fold compared to PelBWT and DsbA, respectively. Based on thioredoxin-1 (TrxA) export studies the triple valine mutant PelB DN5 V3 leader was capable of more efficient cotranslational translocation than PelBWT, but the overall display efficiency improvement over DsbA suggests that besides increased cotranslational translocation other factors contribute to the observed enhancement in DARPin display efficiency.
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