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1. Impact of macromolecular crowding on RNA/spermine complex coacervation and oligonucleotide compartmentalization

   https://doi.org/10.1039/C7SM02146A  

   Marianelli, A. M.; Miller, B. M.; Keating, C. D. (January 2018, Soft Matter)

   We report the effect of neutral macromolecular crowders poly(ethylene glycol) (PEG) (8 kDa) and Ficoll (70 kDa) on liquid–liquid phase separation in a polyuridylic acid (polyU)/spermine complex coacervate system. The addition of PEG decreased both the amount of spermine required for phase separation and the coacervation temperature ( T C ). We interpret these effects on phase behavior as arising due to excluded volume and preferential interactions on both the secondary structure/condensation of spermine-associated polyU molecules and on the association of soluble polyU/spermine polyelectrolyte complexes to form coacervate droplets. Examination of coacervates formed in the presence of fluorescently-labeled PEG or Ficoll crowders indicated that Ficoll is accumulated while PEG is excluded from the coacervate phase, which provides further insight into the differences in phase behavior. Crowding agents impact distribution of a biomolecular solute: partitioning of a fluorescently-labeled U15 RNA oligomer into the polyU/spermine coacervates was increased approximately two-fold by 20 wt% Ficoll 70 kDa and by more than two orders of magnitude by 20 wt% PEG 8 kDa. The volume of the coacervate phase decreased in the presence of crowder relative to a dilute buffer solution. These findings indicate that potential impacts of macromolecular crowding on phase behavior and solute partitioning should be considered in model systems for intracellular membraneless organelles. 

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2. Interfacial stabilization of aqueous two-phase systems: a review

   https://doi.org/10.1039/D3MA00307H  

   Fick, Caitlyn; Khan, Zara; Srivastava, Samanvaya (October 2023, Materials Advances)

   Aqueous two-phase systems (ATPS) are useful in various applications, from purification and separation of biomolecules to wastewater treatment. While they have great utility on their own, there is great interest in discovering how their emulsions, comprising droplets of one aqueous phase dispersed in the other aqueous phase, might be stabilized to enhance their functionality and applications. There are several examples of these systems, but the two most common systems found in the literature are PEG–dextran and complex coacervate ATPS. In this Review, we discuss these systems, their utility, and many different approaches for stabilizing their water/water (w/w) emulsions. We highlight examples wherein interfacial stabilizers such as liposomes, polymers of diverse architectures, colloids of varied shapes and morpholo- gies, and even whole cells have been employed. These stabilization approaches for both PEG–dextran and complex coacervate ATPS are discussed. We conclude with a discussion of the applications of these ATPS and how they can benefit from the creation of corresponding w/w emulsions with stabilized droplets. 

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3. Enhancing pDNA Delivery with Hydroquinine Polymers by Modulating Structure and Composition

   https://doi.org/10.1021/jacsau.3c00126  

   Roy, Punarbasu; Kreofsky, Nicholas W.; Brown, Mary E.; Van Bruggen, Craig; Reineke, Theresa M. (July 2023, JACS Au)

   Quinine is a promising natural product building block for polymer-based nucleic acid delivery vehicles as its structure enables DNA binding through both intercalation and electrostatic interactions. However, studies exploring the potential of quinine-based polymers for nucleic acid delivery applications (transfection) are limited. In this work, we used a hydroquinine-functionalized monomer, HQ, with 2-hydroxyethyl acrylate (HEA) to create a family of 7 polymers (HQ-X, X = mole percentage of HQ) with mole percentages of HQ ranging from 12% to 100%. We developed a flow cytometer-based assay for studying the polymer-pDNA complexes (polyplex particles) directly and demonstrate that polymer composition and monomer structure influence polyplex characteristics such as the pDNA loading and the extent of adsorption of serum proteins on polyplex particles. Biological delivery experiments revealed that maximum transgene expression, outperforming commercial controls, was achieved with HQ-25 and HQ-35 as these two variants sustained gene expression over 96 hours. HQ-44, HQ-60, and HQ-100 were not successful in inducing transgene expression, despite being able to deliver pDNA into the cells, highlighting that the release of pDNA is likely the bottleneck in transfection for polymers with higher HQ content. Using confocal imaging, we quantified the extent of colocalization between pDNA, and lysosomes proving the remarkable endosomal escape capabilities of the HQ-X polymers. Overall, this study demonstrates the advantages of HQ-X polymers as well as provides guiding principles for improving monomer structure and polymer composition, supporting the development of the next generation of polymer-based nucleic acid delivery vehicles harnessing the power of natural products. 

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4. Generation of topologically defined linear and cyclic DNA bottle brush polymers via a graft-to approach

   https://doi.org/10.1039/D5PY00082C  

   Pierini, Nicholas G; Paiva, Wynter A; Durant, Owen C; Dobbins, Aubrianna M; Wheeler, Ben B; Currier, Matthew E; Vesenka, James; Oldenhuis, Nathan J (June 2025, Polymer Chemistry)

   Plasmid-derived DNA bottlebrush polymers with linear and circular topologies are synthesizedviaaqueous grafting of PEG. These constructs exhibit tunable mechanics, enhanced nuclease resistance, and preserve DNA structure and circularity. 

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5. The proto-Nucleic Acid Builder: a software tool for constructing nucleic acid analogs

   https://doi.org/10.1093/nar/gkaa1159  

   Alenaizan, Asem; Barnett, Joshua L; Hud, Nicholas V; Sherrill, C David; Petrov, Anton S (December 2020, Nucleic Acids Research)

   null (Ed.)

   Abstract The helical structures of DNA and RNA were originally revealed by experimental data. Likewise, the development of programs for modeling these natural polymers was guided by known structures. These nucleic acid polymers represent only two members of a potentially vast class of polymers with similar structural features, but that differ from DNA and RNA in the backbone or nucleobases. Xeno nucleic acids (XNAs) incorporate alternative backbones that affect the conformational, chemical, and thermodynamic properties of XNAs. Given the vast chemical space of possible XNAs, computational modeling of alternative nucleic acids can accelerate the search for plausible nucleic acid analogs and guide their rational design. Additionally, a tool for the modeling of nucleic acids could help reveal what nucleic acid polymers may have existed before RNA in the early evolution of life. To aid the development of novel XNA polymers and the search for possible pre-RNA candidates, this article presents the proto-Nucleic Acid Builder (https://github.com/GT-NucleicAcids/pnab), an open-source program for modeling nucleic acid analogs with alternative backbones and nucleobases. The torsion-driven conformation search procedure implemented here predicts structures with good accuracy compared to experimental structures, and correctly demonstrates the correlation between the helical structure and the backbone conformation in DNA and RNA. 

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