Biomedical Engineering Reference
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three peripheral linear strands (P). The two gears share the same C strand but have
different sets of P strands. Each P strand is complement to C strand with a single-
stranded extension acting as foothold. The addition of linker and removal strands
causes the two DNA circles to connect and to continuously roll in one direction
against each other.
The first-generation walkers opened up a new window for DNA molecules to
generate motions with increased complexity, but they have the same shortcoming,
that is, additional DNA strands have to be added manually for every single step of
the walker. This makes it difficult to achieve autonomous and processive motions,
which are important properties for an efficient molecular walker. A number of
systems were developed on the basis of enzymatic catalysis to produce autonomous
motions. Yin et al. reported the first DNA walker that takes advantage of the
activities of ligase and restriction endonuclease [
60
]. The self-assembled footpath
contains anchors at which the walker, a six-nucleotide DNA fragment, can be bound.
In order to take a step, the walker is ligated to the next anchor and then cut from
the previous one by a restriction endonuclease. In this case, each cut destroys the
previous restriction site and each ligation creates a new site so that the walker can
be transferred from anchor to anchor in one direction and cannot move backward.
A similar concept utilizing nicking enzyme to transfer a DNA walker was later
demonstrated by Turberfield and coworkers [
61
]. In this system, the footpath, or
track, is a self-assembled duplex with identical single-stranded stators attached
periodically along its length. The walker is simply a single-stranded DNA that is
able to bind to the stators to form a duplex. A nicking enzyme N.BbvC IB recognizes
the sequence on the walker-stator duplex and cuts the stator to release a short stator
fragment, leaving the walker with a single-stranded overhang free to bind to the
adjacent stator. Once the overhang part of the walker pairs with the next stator, a
simple branch-migration reaction makes the walker to finish the step. Because the
previous stators are cut by the enzyme, the walker cannot take a step backward.
Another DNA walker integrated with DNAzyme was constructed by Tian et al. [
62
]
(Fig.
11.9
c). In this design, the 10-23 DNAzyme plays the roles of both walker
and digesting enzyme which are taken by DNA single strand and nicking enzyme,
respectively, in the last example. Similarly, the footpath is a self-assembled duplex
with periodical stators that is not only complementary to but also the substrate of
the 10-23 DNAzyme. When the DNAzyme binds to one stator, it cuts and releases
the top fragment of the stator. So that the DNAzyme walker is free to bind to the
next stator and accomplish a full step through branch migration.
Several recent researches have made efforts to realize autonomous and processive
walking motion in pure DNA-based systems. These attempts are essentially based
on the principle of DNA hybridization catalysis, which was introduced by Pierce,
Turberfield, as well as Seeman and coworkers [
63
,
64
]. The original purpose
of developing hybridization catalysis mechanism was to enhance the dynamic
hybridization between strands that could form strong secondary structures. As
showninFig.
11.10
a, two hairpin molecules (H1 and H2) have complementary
sequences except for the toehold of H1. They hybridize extremely slowly with each
other because of steric restrictions and stability of the hairpin stems. The addition
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