Dataset Information

MOrPH-PhD: An Integrated Phage Display Platform for the Discovery of Functional Genetically Encoded Peptide Macrocycles.

ABSTRACT: Macrocyclic peptides represent attractive scaffolds for targeting protein-protein interactions, making methods for the diversification and functional selection of these molecules highly valuable for molecular discovery purposes. Here, we report the development of a novel strategy for the generation and high-throughput screening of combinatorial libraries of macrocyclic peptides constrained by a nonreducible thioether bridge. In this system, spontaneous, posttranslational peptide cyclization by means of a cysteine-reactive noncanonical amino acid was integrated with M13 bacteriophage display, enabling the creation of genetically encoded macrocyclic peptide libraries displayed on phage particles. This platform, named MOrPH-PhD, was successfully applied to produce and screen 105- to 108-member libraries of peptide macrocycles against three different protein targets, resulting in the discovery of a high-affinity binder for streptavidin (K D: 20 nM) and potent inhibitors of the therapeutically relevant proteins Kelch-like ECH-associated protein 1 (K D: 40 nM) and Sonic Hedgehog (K D: 550 nM). This work introduces and validates an efficient and general platform for the discovery and evolution of functional, conformationally constrained macrocyclic peptides useful for targeting proteins and protein-mediated interactions.

SUBMITTER: Owens AE

PROVIDER: S-EPMC7099587 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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MOrPH-PhD: An Integrated Phage Display Platform for the Discovery of Functional Genetically Encoded Peptide Macrocycles.

Owens Andrew E AE Iannuzzelli Jacob A JA Gu Yu Y Fasan Rudi R

ACS central science 20200204 3

Macrocyclic peptides represent attractive scaffolds for targeting protein-protein interactions, making methods for the diversification and functional selection of these molecules highly valuable for molecular discovery purposes. Here, we report the development of a novel strategy for the generation and high-throughput screening of combinatorial libraries of macrocyclic peptides constrained by a nonreducible thioether bridge. In this system, spontaneous, posttranslational peptide cyclization by m ...[more]

PMID: 32232137

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Project description:BACKGROUND:Leptospirosis is the most common zoonotic disease worldwide. The diagnostic performance of a serological test for human leptospirosis is mainly influenced by the antigen used in the test assay. An ideal serological test should cover all serovars of pathogenic leptospires with high sensitivity and specificity and use reagents that are relatively inexpensive to produce and can be used in tropical climates. Peptide-based tests fulfil at least the latter two requirements, and ORFeome phage display has been successfully used to identify immunogenic peptides from other pathogens. METHODOLOGY/PRINCIPAL FINDINGS:Two ORFeome phage display libraries of the entire Leptospira spp. genomes from five local strains isolated in Malaysia and seven WHO reference strains were constructed. Subsequently, 18 unique Leptospira peptides were identified in a screen using a pool of sera from patients with acute leptospirosis. Five of these were validated by titration ELISA using different pools of patient or control sera. The diagnostic performance of these five peptides was then assessed against 16 individual sera from patients with acute leptospirosis and 16 healthy donors and was compared to that of two recombinant reference proteins from L. interrogans. This analysis revealed two peptides (SIR16-D1 and SIR16-H1) from the local isolates with good accuracy for the detection of acute leptospirosis (area under the ROC curve: 0.86 and 0.78, respectively; sensitivity: 0.88 and 0.94; specificity: 0.81 and 0.69), which was close to that of the reference proteins LipL32 and Loa22 (area under the ROC curve: 0.91 and 0.80; sensitivity: 0.94 and 0.81; specificity: 0.75 and 0.75). CONCLUSIONS/SIGNIFICANCE:This analysis lends further support for using ORFeome phage display to identify pathogen-associated immunogenic peptides, and it suggests that this technique holds promise for the development of peptide-based diagnostics for leptospirosis and, possibly, of vaccines against this pathogen.

Dataset Information

MOrPH-PhD: An Integrated Phage Display Platform for the Discovery of Functional Genetically Encoded Peptide Macrocycles.

Publications

MOrPH-PhD: An Integrated Phage Display Platform for the Discovery of Functional Genetically Encoded Peptide Macrocycles.

Similar Datasets

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