ABSTRACT
Combinatorial phage display libraries of random peptides can be used to discover the epitopes of antibodies through a procedure termed "biopanning." The affinity isolation of phage-displayed epitope peptidomimetics allows molecular definition of the epitopes of monoclonal antibodies (MAbs). Panels of MAb-specific peptides allow computational prediction of B cell epitopes. Epitope profiles recognized by polyclonal serum samples can also be generated. Detailed step by step protocols and discussion of applications are provided.
Subject(s)
Epitope Mapping/methods , Epitopes, B-Lymphocyte/chemistry , Inovirus , Peptide Fragments/chemistry , Peptide Library , Animals , Antibodies, Monoclonal , Antibody Affinity , Biochemistry , Biomimetics , Computational Biology , Computer Simulation , Epitopes, B-Lymphocyte/immunology , Genetic Engineering , Humans , Molecular Mimicry , Peptide Fragments/immunologyABSTRACT
UNLABELLED: Identifying the epitope to which an antibody binds is central for many immunological applications such as drug design and vaccine development. The Pepitope server is a web-based tool that aims at predicting discontinuous epitopes based on a set of peptides that were affinity-selected against a monoclonal antibody of interest. The server implements three different algorithms for epitope mapping: PepSurf, Mapitope, and a combination of the two. The rationale behind these algorithms is that the set of peptides mimics the genuine epitope in terms of physicochemical properties and spatial organization. When the three-dimensional (3D) structure of the antigen is known, the information in these peptides can be used to computationally infer the corresponding epitope. A user-friendly web interface and a graphical tool that allows viewing the predicted epitopes were developed. Pepitope can also be applied for inferring other types of protein-protein interactions beyond the immunological context, and as a general tool for aligning linear sequences to a 3D structure. AVAILABILITY: http://pepitope.tau.ac.il/
Subject(s)
Algorithms , Epitope Mapping/methods , Peptides/chemistry , Peptides/immunology , Sequence Alignment/methods , Sequence Analysis, Protein/methods , Software , Amino Acid Sequence , Binding Sites , Molecular Sequence Data , Protein BindingABSTRACT
Mapping the epitope of an antibody is of great interest, since it contributes much to our understanding of the mechanisms of molecular recognition and provides the basis for rational vaccine design. Here we present Mapitope, a computer algorithm for epitope mapping. The algorithm input is a set of affinity isolated peptides obtained by screening phage display peptide-libraries with the antibody of interest. The output is usually 1-3 epitope candidates on the surface of the atomic structure of the antigen. We have systematically tested the performance of Mapitope by assessing the effect of the algorithm parameters on the final prediction. Thus, we have examined the effect of the statistical threshold (ST) parameter, relating to the frequency distribution and enrichment of amino acid pairs from the isolated peptides and the D (distance) and E (exposure) parameters which relate to the physical parameters of the antigen. Two model systems were analyzed in which the antibody of interest had previously been co-crystallized with the antigen and thus the epitope is a given. The Mapitope algorithm successfully predicted the epitopes in both models. Accordingly, we formulated a stepwise paradigm for the prediction of discontinuous conformational epitopes using peptides obtained from screening phage display libraries. We applied this paradigm to successfully predict the epitope of the Trastuzumab antibody on the surface of the Her-2/neu receptor in a third model system.
