Multiple alignment and sorting of peptides derived from phage-displayed random peptide libraries with polyclonal sera allows discrimination of relevant phagotopes
Davies, J. M., Scealy, M., Cai, Y., Whisstock, J., Mackay, I. R., & Rowley, M. J. (1999) Multiple alignment and sorting of peptides derived from phage-displayed random peptide libraries with polyclonal sera allows discrimination of relevant phagotopes. Molecular Immunology, 36(10), pp. 659-667.
Biopanning of phage-displayed random peptide libraries is a powerful technique for identifying peptides that mimic epitopes (mimotopes) for monoclonal antibodies (mAbs). However, peptides derived using polyclonal antisera may represent epitopes for a diverse range of antibodies. Hence following screening of phage libraries with polyclonal antisera, including autoimmune disease sera, a procedure is required to distinguish relevant from irrelevant phagotopes. We therefore applied the multiple sequence alignment algorithm PILEUP together with a matrix for scoring amino acid substitutions based on physicochemical properties to generate guide trees depicting relatedness of selected peptides. A random heptapeptide library was biopanned nine times using no selecting antibodies, immunoglobulin G (IgG) from sera of subjects with autoimmune diseases (primary biliary cirrhosis (PBC) and type 1 diabetes) and three murine ascites fluids that contained mAbs to overlapping epitope(s) on the Ross River Virus envelope protein 2. Peptides randomly sampled from the library were distributed throughout the guide tree of the total set of peptides whilst many of the peptides derived in the absence of selecting antibody aligned to a single cluster. Moreover peptides selected by different sources of IgG aligned to separate clusters, each with a different amino acid motif. These alignments were validated by testing all of the 53 phagotopes derived using IgG from PBC sera for reactivity by capture ELISA with antibodies affinity purified on the E2 subunit of the pyruvate dehydrogenase complex (PDC-E2), the major autoantigen in PBC: only those phagotopes that aligned to PBC-associated clusters were reactive. Hence the multiple sequence alignment procedure discriminates relevant from irrelevant phagotopes and thus a major difficulty with biopanning phage-displayed random peptide libraries with polyclonal antibodies is surmounted.
Impact and interest:
Citation counts are sourced monthly from and citation databases.
These databases contain citations from different subsets of available publications and different time periods and thus the citation count from each is usually different. Some works are not in either database and no count is displayed. Scopus includes citations from articles published in 1996 onwards, and Web of Science® generally from 1980 onwards.
Citations counts from theindexing service can be viewed at the linked Google Scholar™ search.
|Item Type:||Journal Article|
|Keywords:||Autoantibodies, Autoimmune disease, Mimotopes, Multiple sequence alignment, Phage display, polyclonal antibody, amino acid substitution, article, bacteriophage, gene cluster, human, human cell, ligand binding, molecular cloning, peptide synthesis, phagocytosis, priority journal, protein conformation, receptor affinity, Algorithms, Antibodies, Viral, Capsid, Capsid Proteins, Diabetes Mellitus, Type 1, Dihydrolipoyllysine-Residue Acetyltransferase, Glutamate Decarboxylase, Humans, Immunoglobulin G, Liver Cirrhosis, Biliary, Peptide Library, Peptides, Pyruvate Dehydrogenase Complex, Sequence Alignment, Viral Envelope Proteins|
|Divisions:||Current > Schools > School of Biomedical Sciences
Current > QUT Faculties and Divisions > Faculty of Health
|Copyright Owner:||Copyright 1999 Elsevier Science Ltd|
|Deposited On:||01 Oct 2015 02:44|
|Last Modified:||01 Oct 2015 02:44|
Repository Staff Only: item control page