Biomedical Engineering Reference
In-Depth Information
behaved significantly different. In the case of boromycin, since it formed a stable
host-guest complex with various amine-based compounds, the long duration block-
age events were in fact attributed to the interaction of the host-guest complex with the
protein pore. Although the un-complexed boromycin also bound to the pore, the
interaction was much weaker, leading to a much shorter duration event. And hence,
the unlabeled boromycin events and the host-guest complex events could be
differentiated based on the residence time although these two types of events had
similar amplitudes. In contrast,
CD bound to the mutant (M113F/K147N)
7
protein
very tightly, so that the channel was partially blocked for a long time after addition of
b
b
CD to the buffer solution. If PMPA or CMPA compounds are additionally added to
the mutant (M113F/K147N)
7
a
HL pore, they will be further captured by the
b
CD
host, thus causing a further current drop (Fig.
13.7b, c
)[
15
].
13.4 Detection of Biomolecules
13.4.1 Peptides
Peptides play important roles in a variety of physiological processes in living
systems. Accordingly, assay of these biologically active molecules are of para-
mount importance in diagnostics and therapeutics [
58
-
60
]. Since the amino acid
components of a peptide can be categorized into four major groups: aromatic,
hydrophobic, positive charge, and negative charge, it is likely that various peptides
may be identified and differentiated by using engineered protein pores with differ-
ent surface functions via weak non-covalent bonding interactions. To demonstrate
this concept, a series of short peptides consisting of mainly aromatic amino acids
and with various lengths was analyzed with a (M113Y)
7
pore, which contains an
aromatic binding site with seven aromatic Tyr side chains. The experimental results
showed that with an increase in the length of the peptide, both the mean residence
time and the current blockage amplitude of the events increased (Fig.
13.8
).
Furthermore, it was found out that the composition of the peptide would also affect
the event residence time and amplitude [
3
]. Take the protein channel modified with
aromatic binding sites for example, aromatic amino acid components of a peptide
contributed more to the residence time and current blockage of the events than other
types of amino acids, although the van der Waals volumes of amino acids also
affected the event signatures. Further experiments showed that with a properly
engineered protein pore, not only could peptides that differed by only an amino acid
be successfully differentiated, but also a mixture of peptides could be simulta-
neously detected (Fig.
13.2
). This weak non-covalent interaction nanopore sensing
approach could even permit differentiation of peptide sequences. For example, a
series of short peptides with the same length and identical composition but with
different sequences such as PYWF, YPWF, YWPF, and YPFW were able to be
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