The Formation and Displacement of Ordered DNA Triplexes in Self-Assembled Three-Dimensional DNA Crystals.

Reconfigurable structures engineered through DNA hybridization and self-assembly offer both structural and dynamic applications in nanotechnology. Here, we have demonstrated that strand displacement of triplex-forming oligonucleotides (TFOs) can be translated to a robust macroscopic DNA crystal by coloring the crystals with covalently attached fluorescent dyes. We show that three different types of triplex strand displacement are feasible within the DNA crystals and the bound TFOs can be removed and/or replaced by (a) changing the pH from 5 to 7, (b) the addition of the Watson-Crick complement to a TFO containing a short toehold, and (c) the addition of a longer TFO that uses the duplex edge as a toehold. We have also proved by X-ray diffraction that the structure of the crystals remains as designed in the presence of the TFOs.

A Rare Case of Aggressive Atypical Cervical Cancer With Multi-Organ Involvement.

Squamous cell carcinoma (SCC) of cervical origin with metastasis to the brain is rare. Our patient was a 30-year-old Caucasian female with squamous cell carcinoma, initially with unknown primary, with metastases to the brain, kidney, cervix, lung, adrenal glands, vulva, pelvic wall, and scalp. She initially presented to her outpatient gynecologist for a vulvar mass. A biopsy of the vulvar mass was consistent with SCC. The patient continued to have fatigue along with thoracic rib pain. An initial work-up was performed, including imaging which showed diffuse metastatic disease involving the lungs, kidneys and adrenal glands, as well as a pathological compression fracture of the seventh thoracic vertebra with cord compression. Brain magnetic resonance imaging (MRI) showed multiple metastatic lesions and she underwent craniotomy for brain lesion resection. Given the aggressive nature of the patient's disease and her symptomatic burden, she was started on chemotherapy in the hospital with Carboplatin, Paclitaxel, and Pembrolizumab.

Fournier's Gangrene in an HIV-Positive Patient on Empagliflozin for the Treatment of Diabetes Mellitus.

Fournier's gangrene is a severe polymicrobial infection that results in necrosis of the perineal and genital fasciae with rapid progression. This case report describes a 55-year-old male with a past medical history of HIV, type 2 diabetes, and hypertension who was diagnosed with Fournier's gangrene after the administration of empagliflozin (Jardiance). The patient presented with a worsening ulcer of the right groin and was diagnosed with Fournier's gangrene based on clinical and radiographic findings. He underwent surgical debridement of the wound. The patient was treated with empiric vancomycin, piperacillin-tazobactam, and clindamycin. Wound cultures grew Streptococcus anginosus and Staphylococcus epidermidis. His antibiotic regimen was simplified to ampicillin-sulbactam. The patient required reconstructive surgery for wound closure after debridement. He received an additional 18 days of augmentin therapy with the resolution of the infectious process. At the time of Fournier's gangrene onset, the patient's last HbA1C level was 8.2%, despite treatment with glipizide and empagliflozin. This case suggests an association between empagliflozin and Fournier's gangrene in the setting of active HIV infection.

Designing Higher Resolution Self-Assembled 3D DNA Crystals via Strand Terminus Modifications.

DNA tensegrity triangles self-assemble into rhombohedral three-dimensional crystals via sticky ended cohesion. Crystals containing two-nucleotide (nt) sticky ends (GA:TC) have been reported previously, and those crystals diffracted to 4.9 Å at beamline NSLS-I-X25. Here, we analyze the effect of varying sticky end lengths and sequences as well as the impact of 5'- and 3'-phosphates on crystal formation and resolution. Tensegrity triangle motifs having 1-, 2-, 3-, and 4-nt sticky ends all form crystals. X-ray diffraction data from the same beamline reveal that the crystal resolution for a 1-nt sticky end (G:C) and a 3-nt sticky end (GAT:ATC) were 3.4 and 4.2 Å, respectively. Resolutions were determined from complete data sets in each case. We also conducted trials that examined every possible combination of 1-nucleotide and 2-nucleotide sticky-ended phosphorylated strands and successfully crystallized all 16 possible combinations of strands. We observed the position of the 5'-phosphate on either the crossover (1), helical (2), or central strand (3) affected the resolution of the self-assembled crystals for the 2-turn monomer (3.0 Å for 1-2P-3P) and 2-turn dimer sticky ended (4.1 Å for 1-2-3P) systems. We have also examined the impact of the identity of the base flanking the sticky ends as well as the use of 3'-phosphate. We conclude that crystal resolution is not a simple consequence of the thermodynamics of the direct nucleotide pairing interactions involved in molecular cohesion in this system.

Self-Assembly of 3D DNA Crystals Containing a Torsionally Stressed Component.

There is an increasing appreciation for structural diversity of DNA that is of interest to both DNA nanotechnology and basic biology. Here, we have explored how DNA responds to torsional stress by building on a previously reported two-turn DNA tensegrity triangle and demonstrating that we could introduce an extra nucleotide pair (np) into the original sequence without affecting assembly and crystallization. The extra np imposes a significant torsional stress, which is accommodated by global changes throughout the B-DNA duplex and the DNA lattice. The work reveals a near-atomic structure of naked DNA under a torsional stress of approximately 14%, and thus provides an example of DNA distortions that occur without a requirement for either an external energy source or the free energy available from protein or drug binding.

A device that operates within a self-assembled 3D DNA crystal.

Structural DNA nanotechnology finds applications in numerous areas, but the construction of objects, 2D and 3D crystalline lattices and devices is prominent among them. Each of these components has been developed individually, and most of them have been combined in pairs. However, to date there are no reports of independent devices contained within 3D crystals. Here we report a three-state 3D device whereby we change the colour of the crystals by diffusing strands that contain dyes in or out of the crystals through the mother-liquor component of the system. Each colouring strand is designed to pair with an extended triangle strand by Watson-Crick base pairing. The arm that contains the dyes is quite flexible, but it is possible to establish the presence of the duplex proximal to the triangle by X-ray crystallography. We modelled the transition between the red and blue states through a simple kinetic model.

An Organic Semiconductor Organized into 3D DNA Arrays by "Bottom-up" Rational Design.

A 3D array of organic semiconductors was assembled using a DNA scaffold. An octameric aniline molecule ("octaniline") was incorporated into a DNA building block based on a dimeric tensegrity triangle. The construct self-assembled to form a 3D crystal. Reversible redox conversion between the pernigraniline and leucoemeraldine states of the octaniline is retained in the crystal. Protonic doping gave emeraldine salt at pH 5, corresponding to the conductive form of polyaniline. Redox cycling within the crystal was visualized by color changes and Raman microscopy. The ease of conversion between the octaniline states suggests that it is a viable electronic switch within a unique 3D structure.

Construction and Structure Determination of a Three-Dimensional DNA Crystal.

Structural DNA nanotechnology combines branched DNA junctions with sticky-ended cohesion to create self-assembling macromolecular architectures. One of the key goals of structural DNA nanotechnology is to construct three-dimensional (3D) crystalline lattices. Here we present a new DNA motif and a strategy that has led to the assembly of a 3D lattice. We have determined the X-ray crystal structures of two related constructs to 3.1 Å resolution using bromine-derivatized crystals. The motif we used employs a five-nucleotide repeating sequence that weaves through a series of two-turn DNA duplexes. The duplexes are tied into a layered structure that is organized and dictated by a concert of four-arm junctions; these in turn assemble into continuous arrays facilitated by sequence-specific sticky-ended cohesion. The 3D X-ray structure of these DNA crystals holds promise for the design of new structural motifs to create programmable 3D DNA lattices with atomic spatial resolution. The two arrays differ by the use of four or six repeats of the five-nucleotide units in the repeating but statistically disordered central strand. In addition, we report a 2D rhombuslike array formed from similar components.