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A structural bioinformatics approach for identifying proteins predisposed to bind linear epitopes on pre-selected target proteins.

ABSTRACT: We have developed a protocol for identifying proteins that are predisposed to bind linear epitopes on target proteins of interest. The protocol searches through the protein database for proteins (scaffolds) that are bound to peptides with sequences similar to accessible, linear epitopes on the target protein. The sequence match is considered more significant if residues calculated to be important in the scaffold-peptide interaction are present in the target epitope. The crystal structure of the scaffold-peptide complex is then used as a template for creating a model of the scaffold bound to the target epitope. This model can then be used in conjunction with sequence optimization algorithms or directed evolution methods to search for scaffold mutations that further increase affinity for the target protein. To test the applicability of this approach we targeted three disease-causing proteins: a tuberculosis virulence factor (TVF), the apical membrane antigen (AMA) from malaria, and hemagglutinin from influenza. In each case the best scoring scaffold was tested, and binders with Kds equal to 37 ?M and 50 nM for TVF and AMA, respectively, were identified. A web server (http://rosettadesign.med.unc.edu/scaffold/) has been created for performing the scaffold search process with user-defined target sequences.

SUBMITTER: Choi EJ

PROVIDER: S-EPMC3601849 | biostudies-literature | 2013 Apr

REPOSITORIES: biostudies-literature

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A structural bioinformatics approach for identifying proteins predisposed to bind linear epitopes on pre-selected target proteins.

Choi Eun Jung EJ Jacak Ron R Kuhlman Brian B

Protein engineering, design & selection : PEDS 20130121 4

We have developed a protocol for identifying proteins that are predisposed to bind linear epitopes on target proteins of interest. The protocol searches through the protein database for proteins (scaffolds) that are bound to peptides with sequences similar to accessible, linear epitopes on the target protein. The sequence match is considered more significant if residues calculated to be important in the scaffold-peptide interaction are present in the target epitope. The crystal structure of the ...[more]

PMID: 23341643

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Project description:BackgroundThe natural antibody responses to B-cell epitopes from dengue structural proteins were assessed using immune sera from people having well-defined past dengue infections with one of the four serotypes.MethodBased on an immune-computational analysis previously conducted, nineteen epitopes from the envelope (E) and eight epitopes from pre-membrane (prM), which were more than 50% conserved across all the four DENV serotypes, were selected. Peptides to represent these B-cell epitopes were obtained from commercially available arrays, and were subjected to enzyme linked immunosorbent assay with sera obtained from dengue seropositive healthy volunteers (DENV1 n = 12: DENV2 n = 12: DENV3 n = 12 and DENV4 n = 12), and 10 dengue seronegative healthy volunteers from Sri Lanka. The cut-off value for the positive antibody response was set by taking the mean response of a peptide to the negative sera plus three standard deviations. The peptides (N = 7) showing the broad immune responses were used to generate antibodies in three mice (Balb/c) batches. The mice antisera were then subjected to microneutralization assays against all the four DENV serotypes. An EC50 viral neutralization ≥ 40 times the serum dilution was considered as neutralizing.ResultsFive of the E-peptide and two prM peptides were recognised by most individuls exposed to infections with each of the four serotypes, showing a serotype cross-reactive broad antibody response. The mice immune sera against the peptides representing the five E protein epitopes neutralized all the four DENV serotypes. Two of these five epitopes are from the Domain II, whereas one of them includes the whole bc-loop region.ConclusionThe antibody responses of highly conserved epitopes across the serotypes, were broadly responsive with sera of all four DENV serotypes collected from individuals infected with only one DENV serotype. Weakly conserved epitopes showed rather specific antibody responses dominated by one or few serotypes.

Dataset Information

A structural bioinformatics approach for identifying proteins predisposed to bind linear epitopes on pre-selected target proteins.

Publications

A structural bioinformatics approach for identifying proteins predisposed to bind linear epitopes on pre-selected target proteins.

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