Herein, we report a DNA nanomachine, built from a DNA-functionalized gold nanoparticle (DNA-AuNP), which moves a DNA walker along a three-dimensional (3-D) DNA-AuNP track and executes the task of releasing payloads. The movement of the DNA walker is powered by a nicking endonuclease that cleaves specific DNA substrates on the track. During the movement, each DNA walker cleaves multiple substrates, resulting in the rapid release of payloads (predesigned DNA sequences and their conjugates). The 3-D DNA nanomachine is highly efficient due to the high local effective concentrations of all DNA components that have been co-conjugated on the same AuNP. Moreover, the activity of the 3-D DNA nanomachine can be controlled by introducing a protecting DNA probe that can hybridize to or dehybridize from the DNA walker in a target-specific manner. This property allows us to tailor the DNA nanomachine into a DNA nanosensor that is able to achieve rapid, isothermal, and homogeneous signal amplification for specific nucleic acids in both buffer and a complicated biomatrix.
Development of an enzyme-powered three dimensional DNA nanomachine for discriminating single nucleotide variants through simulation-guided engineering and noncovalent DNA catalysis.
Herein, we report a bottom-up approach
to assemble a series of
stochastic DNA walkers capable of probing dynamic interactions occurring
at the bio–nano interface. We systematically investigated the
impact of varying interfacial factors, including intramolecular interactions,
orientation, cooperativity, steric effect, multivalence, and binding
hindrance on enzymatic behaviors at the interfaces of spherical nucleic
acids. Our mechanistic study has revealed critical roles of various
interfacial factors that significantly alter molecular binding and
enzymatic behaviors from bulk solutions. The improved understanding
of the bio–nano interface may facilitate better design and
operation of nanoparticle-based biosensors and/or functional devices.
We successfully demonstrate how improved understanding of the bio–nano
interface help rationalize the design of amplifiable biosensors for
nucleic acids and antibodies.
Colloidal
nanoparticle biosensors capable of on-particle biocatalysis
are powerful tools for amplified detection of biomolecules. The development
and practical uses of such concentric amplifiers can be complicated
because of the on-particle biorecognition that involves varying interfacial
factors at the biomolecule–nanoparticle interfaces. Herein,
we reason that a nanoparticle biosensor equipped with an in-solution
biorecognition element may be better fabricated, predicted, controlled,
and performed. The in-solution biorecognition shall also be streamlined
with the on-particle biocatalysis so that the overall analytical and
kinetic performance is not compromised. As a testbed, we introduce
a concentric DNA amplifier driven by an enzyme-powered three-dimensional
DNA nanomachine, where a DNA walker can be instantly assembled onto
a spherical nucleic acid (SNA) track through a polyadenosine anchor.
As such, the free DNA walker can participate in reactions in a homogeneous
solution before assembling to the SNA track. The instant and stable
assembly enabled by both adsorption and complementary base pairing
also ensures rapid on-particle biocatalysis. We demonstrate that the
in-solution biorecognition effectively eliminates the binding hindrance
encountered by the on-particle biorecognition and thus significantly
reduced energy barriers for the detection of nucleic acids and proteins.
Because of the in-solution biorecognition, our system can also be
plugged readily into complex DNA strand displacement networks for
rapid signal amplification.
We have developed an artificial protein scaffold, herewith called a protein vector, which allows linking of an in-vitro synthesised protein to the nucleic acid which encodes it through the process of self-assembly. This protein vector enables the direct physical linkage between a functional protein and its genetic code. The principle is demonstrated using a streptavidin-based protein vector (SAPV) as both a nucleic acid binding pocket and a protein display system. We have shown that functional proteins or protein domains can be produced in vitro and physically linked to their DNA in a single enzymatic reaction. Such self-assembled protein-DNA complexes can be used for protein cloning, the cloning of protein affinity reagents or for the production of proteins which self-assemble on a variety of solid supports. Self-assembly can be utilised for making libraries of protein-DNA complexes or for labelling the protein part of such a complex to a high specific activity by labelling the nucleic acid associated with the protein. In summary, self-assembly offers an opportunity to quickly generate cheap protein affinity reagents, which can also be efficiently labelled, for use in traditional affinity assays or for protein arrays instead of conventional antibodies.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.