In recent years, DNA nanotechnology has emerged as one of the most
powerful strategies for bottom-up construction of nanomaterials. Due to
the high programmability of DNA molecules, their self-assembly can be
rationally designed. Engineered 3D DNA crystals, as critical products
from the design of DNA self-assembly, have been proposed as the
structural scaffolds for organizing nano-objects into three-dimensional,
macroscopic devices. However, for such applications, many obstacles
need to be overcome, including the crystal stability, the
characterization methodology, the revision of crystal designs as well as
the modulation of crystallization kinetics. My PhD research focuses on
solving these problems for engineered 3D DNA crystals to pave the way
for their downstream applications.<br>In this thesis, I started by
enhancing the stability of engineered 3D DNA crystals. I developed a
highly efficient post-assembly modification approach to stabilize DNA
crystals. Enzymatic ligation was performed inside the crystal lattice,
which was designed to covalently link the sticky ends at the crystal
contacts. After ligation, the crystal became a covalently bonded 3D
network of DNA motifs. I investigated the stability of ligated DNA
crystals under a wide range of solution conditions. Experimental data
revealed that ligated DNA crystals had significantly increased
stability. With these highly stabilized DNA crystals, we then
demonstrated their applications in biocatalysis and protein
encapsulation as examples.<br>I also established electron microscope
imaging characterization methods for engineered 3D DNA crystals. For
crystals from large-size DNA motifs, they are difficult to study by
X-ray crystallography because of their limited diffraction resolutions
to no better than 10 Å. Therefore, a direct imaging method by TEM was
set up. DNA crystals were either crushed or controlled to grow into
microcrystals for TEM imaging. To validate the imaging results, we
compared the TEM images with predicted models of the crystal lattice.
With the advance in crystal characterization, DNA crystals of varying
pore size between 5~20 nm were designed, assembled, and validated by TEM
imaging.<br>The post-assembly ligation was further developed to prepare
a series of new materials derived from engineered 3D DNA crystals,
which were inaccessible otherwise. With the directional and spatial
control of ligation in DNA crystal, I prepared new DNA-based materials
including DNA microtubes, complex-architecture crystals, and an
unprecedented reversibly expandable, self-healing DNA crystal. The
integration of weak and strong interactions in crystals enabled a lot of
new opportunities for DNA crystal engineering.<br>In the final chapter,
I investigated the effect of 5’-phosphorylation on DNA crystallization
kinetics. I found that phosphorylation significantly enhanced the
crystallization kinetics, possibly by strengthening the sticky-ended
cohesion. Therefore, DNA crystals can be obtained at much lower ionic
strength after phosphorylation. I also applied the result to controling
the morphology of DNA crystals by tuning the crystallization kinetics
along different crystallographic axes. Together with previously methods
to slow down DNA crystallization, the ability to tune DNA
crystallization kinetics in both ways is essential for DNA crystal
engineering.
| Identifer | oai:union.ndltd.org:purdue.edu/oai:figshare.com:article/7985531 |
| Date | 10 June 2019 |
| Creators | Zhe Li (6581093) |
| Source Sets | Purdue University |
| Detected Language | English |
| Type | Text, Thesis |
| Rights | CC BY 4.0 |
| Relation | https://figshare.com/articles/ENGINEERED_3D_DNA_CRYSTALS_CHARACTERIZATION_STABILIZATION_AND_APPLICATIONS/7985531 |
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