DNA conformations in mismatch repair probed in solution by X-ray scattering from gold nanocrystals.

DNA metabolism and processing frequently require transient or metastable DNA conformations that are biologically important but challenging to characterize. We use gold nanocrystal labels combined with small angle X-ray scattering to develop, test, and apply a method to follow DNA conformations acting in the Escherichia coli mismatch repair (MMR) system in solution. We developed a neutral PEG linker that allowed gold-labeled DNAs to be flash-cooled and stored without degradation in sample quality. The 1,000-fold increased gold nanocrystal scattering vs. DNA enabled investigations at much lower concentrations than otherwise possible to avoid concentration-dependent tetramerization of the MMR initiation enzyme MutS. We analyzed the correlation scattering functions for the nanocrystals to provide higher resolution interparticle distributions not convoluted by the intraparticle distribution. We determined that mispair-containing DNAs were bent more by MutS than complementary sequence DNA (csDNA), did not promote tetramer formation, and allowed MutS conversion to a sliding clamp conformation that eliminated the DNA bends. Addition of second protein responder MutL did not stabilize the MutS-bent forms of DNA. Thus, DNA distortion is only involved at the earliest mispair recognition steps of MMR: MutL does not trap bent DNA conformations, suggesting migrating MutL or MutS/MutL complexes as a conserved feature of MMR. The results promote a mechanism of mismatch DNA bending followed by straightening in initial MutS and MutL responses in MMR. We demonstrate that small angle X-ray scattering with gold labels is an enabling method to examine protein-induced DNA distortions key to the DNA repair, replication, transcription, and packaging.

Direct nanorod assembly using block copolymer-based supramolecules.

Developing routes to control the organization of one-dimensional nanomaterials, such as nanorods, with high precision is critical to generate functional materials since the collective properties depend on their spatial arrangements, interparticle ordering, and macroscopic alignment. We have systematically investigated the coassemblies of nanorods and block copolymer (BCP)-based supramolecules and showed that the energetic contributions from nanorod ligand-polymer interactions, polymer chain deformation, and rod-rod interactions are comparable and can be tailored to disperse nanorods with control over inter-rod ordering and the alignment of nanorods within BCP microdomains. By varying the supramolecular morphology and chemical nature of the nanorods, two highly sought-after morphologies, that is, nanoscopic networks of nanorods and nanorod arrays parallel to cylindrical BCP microdomains can be obtained. The supramolecular approach can be applied to achieve morphological control in nanorod-containing nanocomposites toward fabrication of optical and electronic nanodevices.

Small-molecule-directed nanoparticle assembly towards stimuli-responsive nanocomposites.

Precise control of the spatial organization of nanoscopic building blocks, such as nanoparticles, over multiple length scales is a bottleneck in the 'bottom-up' generation of technologically important materials. Only a few approaches have been shown to achieve nanoparticle assemblies without surface modification. We demonstrate a simple yet versatile approach to produce stimuli-responsive hierarchical assemblies of readily available nanoparticles by combining small molecules and block copolymers. Organization of nanoparticles into one-, two- and three-dimensional arrays with controlled inter-particle separation and ordering is achieved without chemical modification of either the nanoparticles or block copolymers. Nanocomposites responsive to heat and light are demonstrated, where the spatial distribution of the nanoparticles can be varied by exposure to heat or light or changing the local environment. The approach described is applicable to a wide range of nanoparticles and compatible with existing fabrication processes, thereby enabling a non-disruptive approach for the generation of functional devices.

Probing the conformational distributions of subpersistence length DNA.

We have measured the bending elasticity of short double-stranded DNA (dsDNA) chains through small-angle x-ray scattering from solutions of dsDNA-linked dimers of gold nanoparticles. This method, which does not require exertion of external forces or binding to a substrate, reports on the equilibrium distribution of bending fluctuations, not just an average value (as in ensemble fluorescence resonance energy transfer) or an extreme value (as in cyclization), and in principle provides a more robust data set for assessing the suitability of theoretical models. Our experimental results for dsDNA comprising 42-94 basepairs are consistent with a simple wormlike chain model of dsDNA elasticity, whose behavior we have determined from Monte Carlo simulations that explicitly represent nanoparticles and their alkane tethers. A persistence length of 50 nm (150 basepairs) gave a favorable comparison, consistent with the results of single-molecule force-extension experiments on much longer dsDNA chains, but in contrast to recent suggestions of enhanced flexibility at these length scales.

Pyramidal and chiral groupings of gold nanocrystals assembled using DNA scaffolds.

Nanostructures constructed from metal and semiconductor nanocrystals conjugated to and organized by DNA are an emerging class of materials with collective optical properties. We created discrete pyramids of DNA with gold nanocrystals at the tips. By taking small-angle X-ray scattering measurements from solutions of these pyramids, we confirmed that this pyramidal geometry creates structures which are more rigid in solution than linear DNA. We then took advantage of the tetrahedral symmetry to demonstrate construction of chiral nanostructures.

Enzymatic ligation creates discrete multinanoparticle building blocks for self-assembly.

Enzymatic ligation of discrete nanoparticle-DNA conjugates creates nanoparticle dimer and trimer structures in which the nanoparticles are linked by single-stranded DNA, rather than by double-stranded DNA as in previous experiments. Ligation was verified by agarose gel and small-angle X-ray scattering. This capability was utilized in two ways: first, to create a new class of multiparticle building blocks for nanoscale self-assembly and, second, to develop a system that can amplify a population of discrete nanoparticle assemblies.