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Through the NSF Future Manufacturing undergraduate research program at Pasadena City College (PCC), students utilize the tools of synthetic biology to build sustainable, DNA-based materials. The manipulation of DNA enables the construction of microscopic biochemical reactors through the formation of liquid-liquid phase-separated droplets, or DNA condensates. This research investigates the potential of DNA nanostars fused with G-tetraplexes, which can bind hemin, an iron-containing porphyrin co-factor, to form a DNAzyme capable of catalyzing peroxidation reactions within single condensate layers. The in vitro component of this research was enhanced by in silico coarse-grained molecular dynamics simulations, which generated 3D models of the DNA nanostars that allowed student researchers to visualize the behavior of the structures created in the laboratory. Leveraging this computational technique, student researchers developed educational resources and modular lessons to introduce these molecular simulations to a broad student audience at PCC. The simulation programs used, oxDNA and oxView, were instrumental in making this research accessible and engaging for diverse student groups. DNA nanostar simulations were integrated into the General, Organic, and Biochemistry curriculum at PCC, as well as during outreach events such as Girls Science Day, offering students insights into DNA nanostar dynamics and potential applications of DNA-based inventions. This paper details the use of simulation programs to recreate nucleic acid-based nanostructures, advancing the field of DNA nanotechnology. Molecular simulations helped the PCC research students develop experiments that demonstrate how enzymatic activity within DNA droplets can be achieved through G4 complexing. Simulating DNA nanostars with G4s was a profound educational exercise for students, as it taught them about the powerful synergy between in silico and in vitro experimentation. Students also learned about the limitations of modeling biomolecules using computational software, and our G4 simulation results may even inspire the integration of guanine-guanine interactions into the oxDNA program. These findings underscore the significant implications of in silico modeling and structural analysis in biochemical manufacturing and industrial applications, paving the way for further innovations in programmable biomolecular systems. By developing YouTube tutorials that teach students how to carry out nucleic acid simulations on any standard computer, the exploration of DNA dynamics and molecular programming is now widely accessible to both students and educators. 

Abstract Artificial biomolecular condensates are emerging as a versatile approach to organize molecular targets and reactions without the need for lipid membranes. Here we ask whether the temporal response of artificial condensates can be controlled via designed chemical reactions. We address this general question by considering a model problem in which a phase separating component participates in reactions that dynamically activate or deactivate its ability to self-attract. Through a theoretical model we illustrate the transient and equilibrium effects of reactions, linking condensate response and reaction parameters. We experimentally realize our model problem using star-shaped DNA motifs known as nanostars to generate condensates, and we take advantage of strand invasion and displacement reactions to kinetically control the capacity of nanostars to interact. We demonstrate reversible dissolution and growth of DNA condensates in the presence of specific DNA inputs, and we characterize the role of toehold domains, nanostar size, and nanostar valency. Our results will support the development of artificial biomolecular condensates that can adapt to environmental changes with prescribed temporal dynamics. 

Living cells regulate many of their vital functions through dynamic, membraneless compartments that phase separate (condense) in response to different types of stimuli. In synthetic cells, responsive condensates could similarly play a crucial role in sustaining their operations. Here we use DNA nanotechnology to design and characterize artificial condensates that respond to light. These condensates form via the programmable interactions of star-shaped DNA subunits (nanostars), which are engineered to include photo-responsive protection domains. In the absence of UV irradiation, the nanostar interactions are not conducive to the formation of condensates. UV irradiation cleaves the protection domains, increases the nanostar valency and enables condensation. We demonstrate that this approach makes it possible to tune precisely the kinetics of condensate formation by dosing UV exposure time. Our experimental observations are complemented by a computational model that characterizes phase transitions of mixtures of particles of different valency, under changes in the mixture composition and bond interaction energy. In addition, we illustrate how UV activation is a useful tool to control the formation and size of DNA condensates in emulsion droplets, as a prototype organelle in a synthetic cell. This research expands our capacity to remotely control the dynamics of DNA-based components via physical stimuli and is particularly relevant to the development of minimal artificial cells and responsive biomaterials.