Biomedical Engineering Reference
In-Depth Information
For the first time, using a double stranded DNA (34 base pairs) identical to the sequence
of the verotoxin gene as the template molecule, Slinchenko et al. reported the synthesis of
DNA-imprinted polymers which could specifically recognize double-stranded DNA without
the need to destroy the native double-stranded structure of the DNA [74]. It was believed that
the hydrogen bond formation between NH
2
groups of the functional monomer, 2-vinyl-4,6-
diamino-1,3,5-triazine, hydrogen bond acceptors of A-T base pair which are exposed in the
major groove of double-stranded DNA, was the dominate factor for the recognition.
Nishino et al. prepared a molecular imprinted polymer film imprinted with the peptide
sequence from the C-terminus of cytochrome c, alcohol dehydrogenase, and bovine serum
albumen, which was able to selectively capture the target molecules from a mixture solution
of different proteins. They attributed the binding specificity of the epitope-imprinted
polymers, at least in this particular case, to collective hydrogen bonding. When the target
protein contacts a complementary domain, multiple hydrogen bonds are formed between the
oriented groups within the polymer domain and the target protein's C-terminus sequences.
Their results indicated that such a method allowed for the recognition of target proteins based
only on genomic information [75]. Using a 16-residue peptide (lysozyme C, 1.8kDa) as the
template, Brown et al. [76] produced imprinting porous silica scaffolds that could lead to
preferential binding of the whole protein (lysozyme, 14kDa). For comparison, the whole
lysozyme was also imprinted into porous silica scaffolds using sol-gel processing. Both
imprinting methods are presented in Figure 6. After removing template, both kinds of
scaffolds were exposed to lysozyme and/or RNase A, which was used as a competitor
molecule of comparable size. When comparing whole protein- to peptide-imprinted scaffolds,
similar amounts of lysozyme and RNase were bound from single protein solutions. However,
while whole lysozyme-imprinted scaffolds showed about 4:1 preferential binding of lysozyme
to RNase, peptide-imprinted scaffolds failed to show statistical significance, even though a
slight preferential binding trend was present. It is suggested that there is potential for further
development of the epitope approach to improve selectivity.
Actually, concerning protein recognition, it is difficult to evaluate the true efficiency of
the epitope approach. In most experiments reported by far, the epitope exploited were up to
few amino acid long and the template they were intended to mimic were just small peptides,
only 8-10 amino acid long, thus non-three-dimensionally structured. Concerning the
exploitation of the epitope approach for the recognition a protein (sequences above 200 amino
acids), attention should be paid to three-dimensional structure of the selected epitopes, as it
would play a significant role in recognition.
5. Mechanism of the Molecular Imprinting and Recognition
It is well known now that molecular imprinting generally involves arranging
polymerizable functional monomers around a template, followed by polymerization and
template removal. The quality and the performance of the molecular imprinted polymers
depend on carefully tailoring of the physical and chemical nature of the monomers and the
interactions between them, the polymerization and its affect on the porous structure, and the
rebinding ability of the imprinted cavities. Ensuring that the functional groups of the template
molecule are interacting with complementary functionality of polymer-forming compounds is
the major factor in this technique.
