Designing chemical reaction networks for self-regulating colloidal assemblies and DNA delivery applications
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Abstract
Biological self-assemblies for e.g., microtubules regulate in space and time via kinetically controlled reaction networks, feedback loops, and energy dissipation giving rise to non-equilibrium processes. Such structures provide an interesting approach for the development of artificial self-assembling molecular systems and materials functions. Early examples of synthetic out-of-equilibrium self-assemblies primarily include supramolecular systems that form transient gels, polymers, vesicles, and micelles. Colloids are rarely used as building blocks. However, they can give rise to functional materials with unique catalytic, photonic, magnetic, and electronic properties originating not only from the positional or dimensional order of the building blocks, but also from the type of material bulk (metallic, polymeric, semiconductor, inorganic). Employing biocompatible material for their fabrication can even pave the way into biomedical applications such as drug delivery and diagnostics. They can even serve as model systems to elucidate physical concepts underlying out-of-equilibrium biological self-assemblies.
The research of this thesis aims at developing new concepts for transient colloidal co-assemblies by either coupling or integrating micron-sized colloidal particles to chemical reaction networks (CRNs) of different origins. CRNs are composed of antagonistic activation and deactivation pathways decoupled by either chemical modulation, temporal modulation, or by energy dissipation strategy (reminiscent of biological self-assemblies). The activation pathway induces the formation of assembled structures that are subsequently degraded by the deactivation pathway giving the transient structures a limited lifetime. The kinetics of the antagonistic pathways are tunable and can be tuned to further modulate the lifetime and structural properties of the transient assemblies. Two major challenges addressed in this thesis to achieve successful transient assembly of colloids are: (i) time-dependency of interparticle potentials and (ii) matching the dynamics of micron-sized particles with kinetics of CRNs. Both conditions are indispensable to target transient structures without the system falling into kinetic traps.
Within the framework of this thesis, three main design strategies are established to program transient co-assemblies of micron-scale colloids.
In the first part of the thesis, a pair of pH-responsive, hetero-complementary colloids are integrated within a pH-feedback system (pH-FS) wherein an alkaline trigger facilitates transient assemblies and autonomous acid formation via an enzymatic cascade subsequent leads to disassembly. The system features advanced chemo-structural feedback where co-assemblies (structures) accelerate their own destruction (negative feedback).
To increase the programmability and modularity of transient assemblies, the second part of the thesis makes use of DNA-based networks such as toehold-mediated DNA strand displacement (TMDSD) reaction networks. A TMDSD reaction cascade directs two different colloids into transient co-assemblies. The colloids are functionalized with DNA and become an integral part of the network. The particles transduce two orthogonally different DNA trigger strands to transiently introduce a linker which brings two colloids together into co-assemblies. The system follows a complex trajectory passing through a transient state in the middle and ultimately reaching thermodynamic equilibrium. Although dynamic, the system operates under non-dissipative conditions as it resides in a new state (lowest energy state) and the original state cannot be acquired. The modularity of the design allows the installation of delay phases and accelerators by interconnecting modules to the upstream and downstream of the core network.
Moving away from the thermodynamically driven classic TMDSD reactions, the third part of the thesis deals with ATP-fueled enzymatic TMDSD reaction cascade which operates under dissipative conditions. The ATP-powered ligation and restriction of DNA components transiently generates a linker strand at the molecular level which temporally controls the downstream co-assembly of colloids. The resetting ability of the network restores the original state of the system and allows the system to be reused for subsequent cycles. Because of their high robustness, ATP-fueled reaction networks are also installed in the extracellular medium of HeLa cells (cancer cells) to develop a DNA delivery system. The delivery mechanism proposed holds potential applications in gene therapy and gene silencing where therapeutic oligonucleotides such as small interfering RNA (siRNA) and micro RNA (miRNA) can be selectively delivered to target cells and tissues in response to ATP.
In a future perspective, strategies developed in this thesis for the fabrication of transient colloidal assemblies represent a step towards functional materials. The design principles can be extended to colloids of different origins, depending on the type of material or application they are targeted at. The modularity of CRNs. especially DNA-based networks, allows the insertion of additional feedback loops to achieve advance dynamic functionalities such as oscillations and bistability.