Molecular recognition of damaged DNA using synthetic affinity reagents
Potentially carcinogenic DNA damage that may be derived from dietary sources can
be quantified using a number of methods. Some of the best of these methods are
based on the use of antibodies that have high affinity and specificity for modified
DNA bases. However, given the sometimes limited availability of antibodies there is
a requirement for novel reagents that mimic the properties of the best antibodies.
From a chemical perspective antibodies are `over engineered' and synthetic affinity
reagents would recreate the optimal properties of the binding site with respect to
selectivity and affinity.
Phage display was used to identify amino acid residues that contribute to recognition
of a diet derived DNA adduct 06-carboxymethyl-2'-deoxyguanosine. However, this
information was not sufficient to synthesise a novel peptide reagent directly and it
was decided to develop an approach whereby a library of compounds was synthesised
which includes the characteristics expected within the binding site of an antibody.
This library was based around cholic acid, a readily available scaffold molecule with
suitable functionality compatible with linking amino acids and a fluorescent tag.
A synthetic procedure based on the formation of stable oxime derivatives of cholic
acid was optimised and a small library of pyrene-tagged trisubstituted cholic acid
derivatives was characterised. The library was examined for its ability to bind to
affinity columns containing immobilised 06-carboxymethylguanosine and there was
some evidence that indicated specific binding of a subset of library molecules