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Epitope mapping and key amino acid identification of anti-CD22 immunotoxin CAT-8015 using hybrid -lactamase display

2010; Oxford University Press; Volume: 24; Issue: 4 Linguagem: Inglês

10.1093/protein/gzq114

1741-0134

David Bannister, Bojana Popovic, Sridha Sridharan, F. Giannotta, Patrice Filée, Nursel Yilmaz, Ralph Minter,

Bacteriophages and microbial interactions

Monoclonal antibodies are a commercially successful class of drug molecules and there are now a growing number of antibodies coupled to toxic payloads, which demonstrate clinical efficacy. Determining the precise epitope of therapeutic antibodies is beneficial in understanding the structure–activity relationship of the drug, but in many cases is not done due to the structural complexity of, in particular, conformational protein epitopes. Using the immunotoxin CAT-8015 as a test case, this study demonstrates that a new methodology, hybrid β-lactamase display, can be employed to elucidate a complex epitope on CD22. Following insertion of random CD22 gene fragments into a permissive site within β-lactamase, proteins expressed in Escherichia coli were first screened for correct folding by resistance to ampicillin and then selected by phage display for affinity to CAT-8015. The optimal protein region recognised by CAT-8015 could then be used as a tool for fine epitope mapping, using alanine-scanning analysis, demonstrating that this technology is well suited to the rapid characterisation of antibody epitopes.