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Compositional bias in naïve and chemically-modified phage-displayed libraries uncovered by paired-end deep sequencing

Abstract : Understanding the composition of a genetically-encoded (GE) library is instrumental to the success of ligand discovery. In this manuscript, we investigate the bias in GE-libraries of linear, macrocyclic and chemically post-translationally modified (cPTM) tetrapeptides displayed on the M13KE platform, which are produced via trinucleotide cassette synthesis (19 codons) and NNK-randomized codon. Differential enrichment of synthetic DNA {S}, ligated vector {L} (extension and ligation of synthetic DNA into the vector), naïve libraries {N} (transformation of the ligated vector into the bacteria followed by expression of the library for 4.5 hours to yield a "naïve" library), and libraries chemically modified by aldehyde ligation and cysteine macrocyclization {M} characterized by paired-end deep sequencing, detected a significant drop in diversity in {L} → {N}, but only a minor compositional difference in {S} → {L} and {N} → {M}. Libraries expressed at the N-terminus of phage protein pIII censored positively charged amino acids Arg and Lys; libraries expressed between pIII domains N1 and N2 overcame Arg/Lys-censorship but introduced new bias towards Gly and Ser. Interrogation of biases arising from cPTM by aldehyde ligation and cysteine macrocyclization unveiled censorship of sequences with Ser/Phe. Analogous analysis can be used to explore library diversity in new display platforms and optimize cPTM of these libraries.
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https://hal.inrae.fr/hal-02623801
Déposant : Migration Prodinra <>
Soumis le : mardi 26 mai 2020 - 09:25:06
Dernière modification le : vendredi 12 juin 2020 - 10:43:26

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Bifang He, Katrina F. Tjhung, Nicholas J. Bennett, Ying Chou, Andrea Rau, et al.. Compositional bias in naïve and chemically-modified phage-displayed libraries uncovered by paired-end deep sequencing. Scientific Reports, Nature Publishing Group, 2018, 8 (1214), ⟨10.1038/s41598-018-19439-2⟩. ⟨hal-02623801⟩

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