Towards Integrated Molecular Machines: Structural, Mechanical, and Computational Motifs

Citation

Sarraf, Namita (2023) Towards Integrated Molecular Machines: Structural, Mechanical, and Computational Motifs. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/cdwp-c709. https://resolver.caltech.edu/CaltechTHESIS:01272023-184413283

Abstract

The programmability of DNA has made it well-suited for building molecular machines, performing nanoscale self-assembly, and computing via biochemical circuits. In the last few decades, great strides have been made in characterizing the interactions between DNA molecules such that they can be predicted and engineered.

The development of frameworks for those interactions has enabled the construction of more complex molecular systems that can execute specified programs. Such programs have included mechanical tasks, like walking and sorting cargo; assembly and reconfiguration of 2D and 3D shapes; and computation, like Boolean logic and pattern recognition.

However, the continuing development of more complex molecular programs relies upon expanding the modules available for molecular systems to use to execute them. Expanded functionality of mechanical, structural, and computation modules are required in order to build compound systems that can interact with the physical world, reconfigure, and analyze signals in a variety of interesting ways. In this dissertation, we will discuss our contributions to this effort, which include exploring a motif for molecular robotic behavior, characterizing tile-tile interactions, and developing new capabilities for bimolecular circuits.

Within the framework of a maze-solving molecular robot, we aim to implement walking behavior on DNA origami that introduces a surface modification via a four-way strand displacement reaction. Surprisingly, our experiments suggest that the walking behavior is at least two orders of magnitude slower than expected. To understand why, we quantitatively explore to what extent the speed and completion level of the robot can be modulated by design considerations such as toehold lengths, track redundancy, and strand purity. Another factor affecting the reaction rate is the number of tethering points, and we demonstrate an order of magnitude speed up in the four-way strand displacement reaction when we remove one tethering point. The characterization of a surface-modifying four-way strand displacement reaction is a useful tool for the continued development of molecular robots with more complex functionality.

Free-floating DNA origami tiles, called invaders here, can swap out DNA origami tiles within larger assemblies via a technique called tile displacement, which has previously been demonstrated using single tile and dimer invaders with 4- and 9-tile arrays. We introduce initial structures and invading assemblies with more complex shapes. We explore the robustness of this reaction by testing a variety of edge configurations and comparing their reaction rates. We demonstrate tunable growth of one of the invaders, which can grow into polymers of arbitrary length or close into 3D structures. By a tile displacement reaction, we reconfigure the 3D structures into 2D. The invaders with complex shapes are able to reconfigure the original tile assembly at rates comparable to simpler tile displacement reactions, and two reconfiguration events can take place sequentially or simultaneously.

Finally, we build two new modules for use with biochemical circuits. The first, a loser-take-all circuit, yields binary outputs indicating which analog signal is the smallest among all inputs. We implement a signal reversal function that converts the smallest input to the largest output, which can then be composed with a previously developed winner-take-all function to achieve loser-take-all. By making concentration adjustments, we can mitigate biases in the circuit that are a result of sequence-dependent different in reaction rates. We experimentally demonstrate a three-input loser-take-all circuit with nine input combinations. With further development, this circuit could be used to implement the activation function in neural networks that perform pattern classification according to which memory an input pattern is least similar to.

The second circuit processes information using temporary memory. We design and implement a circuit that outputs distinct logic decisions based on relative timing information of a pair inputs and their logic values. We show that we can mitigate crosstalk in the circuit by utilizing mismatches and adjusting toehold lengths. The circuit is able to display clear ON-OFF separation at time intervals as short as one minute between the two inputs arriving.

Thesis Files