Rapid and cost-effective epitope mapping using PURE ribosome display coupled with next-generation sequencing and bioinformatics

A novel, efficient and cost-effective approach for epitope identification of an antibody has been developed using a ribosome display platform. This platform, known as PURE ribosome display, utilizes an Escherichia coli-based reconstituted cell-free protein synthesis system (PURE system). It stabiliz...

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Veröffentlicht in:Journal of bioscience and bioengineering Jg. 137; H. 4; S. 321 - 328
Hauptverfasser: Jia, Beixi, Ojima-Kato, Teruyo, Kojima, Takaaki, Nakano, Hideo
Format: Journal Article
Sprache:Englisch
Veröffentlicht: Japan Elsevier B.V 01.04.2024
Schlagworte:
algorithms
Anti-HA tag antibody
bioinformatics
Cell-free protein synthesis
cost effectiveness
Epitope mapping
epitopes
hemagglutinins
monoclonal antibodies
oligonucleotides
peptide libraries
Python program
Ribosome display
ribosomes
Epitope mapping
Anti-HA tag antibody
Python program
Cell-free protein synthesis
Ribosome display
ISSN:1389-1723, 1347-4421, 1347-4421
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Zusammenfassung:A novel, efficient and cost-effective approach for epitope identification of an antibody has been developed using a ribosome display platform. This platform, known as PURE ribosome display, utilizes an Escherichia coli-based reconstituted cell-free protein synthesis system (PURE system). It stabilizes the mRNA-ribosome-peptide complex via a ribosome-arrest peptide sequence. This system was complemented by next-generation sequencing (NGS) and an algorithm for analyzing binding epitopes. To showcase the effectiveness of this method, selection conditions were refined using the anti-PA tag monoclonal antibody with the PA tag peptide as a model. Subsequently, a random peptide library was constructed using 10 NNK triplet oligonucleotides via the PURE ribosome display. The resulting random peptide library-ribosome-mRNA complex was selected using a commercially available anti-HA (YPYDVPDYA) tag monoclonal antibody, followed by NGS and bioinformatic analysis. Our approach successfully identified the DVPDY sequence as an epitope within the hemagglutinin amino acid sequence, which was then experimentally validated. This platform provided a valuable tool for investigating continuous epitopes in antibodies.
Bibliographie:ObjectType-Article-1
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ISSN:1389-1723
1347-4421
1347-4421
DOI:10.1016/j.jbiosc.2024.01.008