Abstract
Nucleic acid-based technologies are an emerging research focus area for pharmacological and biological studies because they are biocompatible and can be designed to produce a variety of scaffolds at the nanometer scale. The use of nucleic acids (ribonucleic acid (RNA) and/or deoxyribonucleic acid (DNA)) as building materials in programming the assemblies and their further functionalization has recently established a new exciting field of RNA and DNA nanotechnology, which have both already produced a variety of different functional nanostructures and nanodevices. It is evident that the resultant architectures require detailed structural and functional characterization and that a variety of technical approaches must be employed to promote the development of the emerging fields. Small-angle X-ray and neutron scattering (SAS) are structural characterization techniques that are well placed to determine the conformation of nucleic acid nanoparticles (NANPs) under varying solution conditions, thus allowing for the optimization of their design. SAS experiments provide information on the overall shapes and particle dimensions of macromolecules and are ideal for following conformational changes of the molecular ensemble as it behaves in solution. In addition, the inherent differences in the neutron scattering of nucleic acids, lipids, and proteins, as well as the different neutron scattering properties of the isotopes of hydrogen, combined with the ability to uniformly label biological macromolecules with deuterium, allow one to characterize the conformations and relative dispositions of the individual components within an assembly of biomolecules. This article will review the application of SAS methods and provide a summary of their successful utilization in the emerging field of NANP technology to date, as well as share our vision on its use in complementing a broad suite of structural characterization tools with some simulated results that have never been shared before.
Highlights
Nucleic acid-based nanoparticles (NANPs) [1,2,3,4,5,6,7,8,9,10,11,12,13] and other nucleic acid-based nanodevices [14,15,16,17] are an emerging research focus area in pharmacological and biological studies
None of the aforementioned techniques allows for direct visualization of large (>100 kDa) three-dimensional NANPs in solution, and they often require working with high concentrations of NANPs
Small-angle X-ray and neutron scattering (SAS) was first described for biomolecules in the late 1940s [43] and has been widely used for several decades to solve problems requiring an understanding of the nanoscale phenomena
Summary
Nucleic acid-based nanoparticles (NANPs) [1,2,3,4,5,6,7,8,9,10,11,12,13] and other nucleic acid-based nanodevices [14,15,16,17] are an emerging research focus area in pharmacological and biological studies. Conventional characterization techniques include the routinely used analysis of NANPs by native-PAGE (non-denaturing PolyAcrylamide Gel Electrophoresis) [23], dynamic light scattering (DLS) [4], atomic force microscopy (AFM) [24], and more sophisticated methods employing cryogenic-electron microscopy (cryo-EM) [25], nuclear magnetic resonance (NMR) [26], and X-ray crystallography [27] (see Table 1 for a comparison of the advantages and limitations of these structural characterization techniques). Size distributions, shape Sample preparation is relatively simple Structure in native state. Size distributions, shape Produces high resolution images that can provide information about structure and elemental composition High resolution TEM has Å resolution. Sample volumes are small (μL) Particle size across a broad range (~0.1 nm to ~10 μm) Allows measurements under physiological conditions
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