Page 67 - ASME_NEMB_2016_Program

TRACK 5 TRACK 5 Technical Program

crowding is known to affect molecular motor transport, enzyme function, and Jacob Berlin, City of Hope, Beckman Research Inst., Duarte, CA,
other molecular interactions. United States

4:40pm Regulation-at-a-Distance of Biomolecular Interactions The controlled assembly of biocompatible nanoparticle aggregates using
Using a DNA Origami Nanoactuator small molecule crosslinker has been a long standing challenge, likely ow-
ing to difficulties in controlling rates of initiation, propagation and termina-
Technical Presentation. NEMB2016-5954 tion. Here we demonstrate that adjusting the concentration of the starting
nanoparticles or the crosslinker allows for the preparation of relatively
homogenous aggregates from metallic nanoparticles of varied composition
Yonggang Ke, Emory School of Medicine, Atlanta, GA, United and size, presumably by controlling the rates of initiation and propogation.
States, Travis Meyer, Emory University/Georgia Institute of Technol- Capping reactive thiols on the formed aggregates with PEG-maleimide
ogy, Atlanta, GA, United States provides a termination step and renders the aggregates stable and biocom-
patible. The size of the aggregates can be systematically adjusted. The ag-
The creation of nanometer-sized structures that exhibit controllable mo- gregates are biocompatible and show no toxicity when incubated with cells.
tions and functions is a critical step towards building nanomachines. Recent Finally, the aggregates are highly stable and appear unchanged after uptake
developments in the field of DNA nanotechnology have begun to address by cells. It is expected that this straightforward and inexpensive assembly of
these goals, demonstrating complex static or dynamic nanostructures made highly stable nanoparticle aggregates will expand the biological applications
of DNA. Here we have designed and constructed a rhombus-shaped DNA of this class of materials. Furthermore, this method for preparing aggregates
origami “nanoactuator” that uses mechanical linkages to copy distance is highly modular as the crosslinker, the building block nanoparticles and the
changes induced on one half (“the driver”) to be propagated to the other exterior coating can all be independently varied and the use of alternative
half (“the mirror”). We demonstrated the ability to control this dynamic DNA crosslinkers and capping agents will enable applications in diverse material
origami with long-range allosteric activation properties, wherein the binding applications.
of an effector molecule controls its global shape. By combining this nano-
actuator with split Enhanced Green Fluorescent Protein (eGFP), we have 5:20pm Heat-Shrunken Hierarchical Silica Nanomembrane for
constructed a DNA-protein hybrid nanostructure that demonstrates tunable Solid Phase DNA Extraction
fluorescent behaviors via long-range allosteric regulation. In addition, the na-
noactuator can be used as a sensor that responds to specific stimuli, includ-
ing changes in buffer composition and the presence of restriction enzymes Technical Presentation. NEMB2016-6013
or specific nucleic acids.
Ye Zhang, Johns Hopkins University, Baltimore, MD, United States,
We expect the same mechanism can potentially be applied to other pro- Yi Zhang, Institute of Bioengineering and Nanotechnology, Singa-
teins, small molecules, and nanoparticles for constructing DNA-controlled ar- pore,Singapore, Kelvin Liu, Circulomics Inc., Baltimore, MD, United
tificial nanodevices with tunable enzymatic, photonic, or plasmonic functions. States, Tza-huei Wang, Johns Hopkins University, Baltimore, MD,
In addition, the nanodevice can serve as a general platform for studying United States
weak molecular interactions at the single-molecule level (for instance, weak
protein-protein interactions). Single-molecule experiments are generally Introduction: The ability to obtain large quantities of high molecular weight
limited to strongly interacting pairs because the short binding time between and high purity DNA is of rising importance as genetic analysis tools
weak pairs often make it impossible to monitor. In addition, single-molecule become increasingly sophisticated. Conventionally, liquid phase DNA
methods typically require low concentrations of molecules, which further extraction techniques, such as Phenol/Chloroform precipitation, are used
reduce the frequency of weak molecular interactions. Our device provides for high quality sample preparation; however, this approach is laborious,
a potential solution for observing these weak interactions: by increasing time-consuming and highly operator-dependent. Solid phase DNA extraction
the local concentration of molecules and by providing rationally designed via silica adsorption, such as spin columns and magnetic particles, are gain-
spatial confinement, we may enhance the weak interactions to enable moni- ing popularity these days due to their speed, ease of use, and generally
toring such events at the single-molecule level.
high performance. However, they tend to induce shear forces that fragment
DNA and reduce DNA quality. In this paper, we present a simple strategy to
This novel mechanism could also potentially enable construction a group create a high surface area silica nanomembrane that contains a high density
of sensors that respond to a wide range of stimuli besides what have been of hierarchical micro- and nanoscale features. This material is created by de-
demonstrated in this work. In these sensors, the sequences of the detect- positing silica onto a thermoplastic polyolefin (PO) film and heat-shrinking in
ing strands were designed independently without regard to the underlying an oven. The resultant membrane exhibits overlaying hierarchical structures
DNA origami structures that carried the fluorescence reporters. Therefore, from nano to micro scale, which can be fine-tuned through silica deposition
these sequences can be modified to enable the DNA origami nanodevice thickness. These nanostructures significantly increase the total surface area
to sense other types of stimuli ranging from small molecules to large protein of silica on the membrane with exceptional adsorption capacity for DNA as
complexes. For instance, implementing aptamer sequences can potentially a novel substrate for solid phase extraction. DNA isolated with the proposed
allow the device to detect specific proteins.
nanomembrane shows higher recovery yield and better integrity comparing
with commercial columns and particles, as well as comparable performance
Finally, the nanodevice could be adapted to serve as a quantitative platform with gold standard Phenol/Chloroform precipitation.
for a low-cost electrophoretic mobility shift assay. The allosteric mechanism
and the large size of the origami devices are two of the advantages for stim- Materials and Methods: The nanomembrane was fabricated by depositing
uli detection using this technique. Alternatively, the drastic conformational silica onto both sides of the PO film using electron beam physical vapor
change of the device in presence of specific targets can be visually detect- deposition. Then the silica-coated film was baked at 300°F for 3 min to
ed at the single-molecule level through atomic force microscopy or electron induce surface wrinkling. The resulted shrunk film is smaller than 10% of its
microscopy. For fluorescence-based detection, the three-dimensional shape original size and exhibits hierarchical nanostructures under scanning elec-
and the chemical addressability of the origami device allows for the incorpo- tron microscope. Nucleic acid extraction can then be performed using a
ration of several dye/quencher pairs along the arms of the device, thereby standard bind, wash, and elute protocol in as little as 45 minutes.
making read-out of the nanoswitches more sensitive.
Results and Discussion: The nanomembrane surface topography can be
fine-tuned by varying the thickness of silica deposition. With 2nm silica, the
5:00pm Controlled Assembly Of Biocompatible Metallic Nano- membrane displays only micro-scale ridges. With 20nm and 50nm silica, na-
aggregates Using A Small Molecule Crosslinker noscale wrinkles and flakes emerge and interweave with micro-ridges, form-
ing overlaying hierarchical nanostructures. Once the silica layer exceeds 67
Technical Presentation. NEMB2016-6087 100nm, a high density of nano-scale flakes interweave to form secondary