Purification of anti-MUC1 antibodies by peptide mimotope affinity chromatography using peptides derived from a polyvalent phage display library.
A polyvalent, lytic phage display system (T7Select415-1b) displaying a random peptide library has been investigated for its ability to discover novel mimotopes reactive with the therapeutic monoclonal antibody C595. Sequence analysis of enriched phage lead to the identification of a predominant sequence RNREAPRGKICS, and two other consensus sequences RXXP and RXP. The novel synthetic peptide RNREAPRGKICS was linked to beaded agarose and the performance as a mimotope affinity chromatography matrix evaluated. Antibody purified using the novel matrix was found to be of higher specific reactivity than antibody purified using the conventional epitope matrix (peptide APDTRPAPG). The RNREAPRGKICS peptide binding to C595 demonstrated a higher equilibrium association constant (K(A)=0.75 x 10(6)) than the epitope peptide (K(A)=0.16 x 10(6)). Circular dichroism showed that the novel peptide had a more highly ordered structure at 4 degrees C and room temperature, than the epitope peptide.
Smith
Richard G
RG
University of Nottingham, Cancer Research Laboratories, School of Pharmaceutical Sciences, UK. rgsmith@dstl.gov.uk
Missailidis
Sotiris
S
Price
Michael R
MR
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000302:isolation & purification*; D001435:Bacteriophages / Q000235:genetics; D001483:Base Sequence; D002846:Chromatography, Affinity / Q000379:methods*; D002942:Circular Dichroism; D018716:Molecular Mimicry; D008969:Molecular Sequence Data; D018396:Mucin-1 / Q000276:immunology*; D010455:Peptides / Q000737:chemistry / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D013050:Spectrometry, Fluorescence
D000911:Antibodies, Monoclonal; D018396:Mucin-1; D010455:Peptides