Two computational primitives for algorithmic self-assembly: Copying and counting

Copying and counting are useful primitive operations for computation and construction. We have made DNA crystals that copy and crystals that count as they grow. For counting, 16 oligonucleotides assemble into four DNA Wang tiles that subsequently crystallize on a polymeric nucleating scaffold strand, arranging themselves in a binary counting pattern that could serve as a template for a molecular electronic demultiplexing circuit. Although the yield of counting crystals is low, and per-tile error rates in such crystals is roughly 10%, this work demonstrates the potential of algorithmic self-assembly to create complex nanoscale patterns of technological interest. A subset of the tiles for counting form information-bearing DNA tubes that copy bit strings from layer to layer along their length

Modeling DNA beacons at the mesoscopic scale

We report model calculations on DNA single strands which describe the equilibrium dynamics and kinetics of hairpin formation and melting. Modeling is at the level of single bases. Strand rigidity is described in terms of simple polymer models; alternative calculations performed using the freely rotating chain and the discrete Kratky-Porod models are reported. Stem formation is modeled according to the Peyrard-Bishop-Dauxois Hamiltonian. The kinetics of opening and closing is described in terms of a diffusion-controlled motion in an effective free energy landscape. Melting profiles, dependence of melting temperature on loop length, and kinetic time scales are in semiquantitative agreement with experimental data obtained from fluorescent DNA beacons forming poly(T) loops. Variation in strand rigidity is not sufficient to account for the large activation enthalpy of closing and the strong loop length dependence observed in hairpins forming poly(A) loops. Implications for modeling single strands of DNA or RNA are discussed.Comment: 15 pages, 17 figures, submitted to Eur. J. Phys.

Capturing the essence of folding and functions of biomolecules using Coarse-Grained Models

The distances over which biological molecules and their complexes can function range from a few nanometres, in the case of folded structures, to millimetres, for example during chromosome organization. Describing phenomena that cover such diverse length, and also time scales, requires models that capture the underlying physics for the particular length scale of interest. Theoretical ideas, in particular, concepts from polymer physics, have guided the development of coarse-grained models to study folding of DNA, RNA, and proteins. More recently, such models and their variants have been applied to the functions of biological nanomachines. Simulations using coarse-grained models are now poised to address a wide range of problems in biology.Comment: 37 pages, 8 figure

Algorithmic Self-Assembly of DNA: Theoretical Motivations and 2D Assembly Experiments

Biology makes things far smaller and more complex than anything produced by human engineering. The biotechnology revolution has for the first time given us the tools necessary to consider engineering on the molecular level. Research in DNA computation, launched by Len Adleman, has opened the door for experimental study of programmable biochemical reactions. Here we focus on a single biochemical mechanism, the self-assembly of DNA structures, that is theoretically sufficient for Turing-universal computation. The theory combines Hao Wang?s purely mathematical Tiling Problem with the branched DNA constructions of Ned Seeman. In the context of mathematical logic, Wang showed how jigsaw-shaped tiles can be designed to simulate the operation of any Turing Machine. For a biochemical implementation, we will need molecular Wang tiles. DNA molecular structures and intermolecular interactions are particularly amenable to design and are sufficient for the creation of complex molecular objects. The structure of individual molecules can be designed by maximizing desired and minimizing undesired Watson-Crick complementarity. Intermolecular interactions are programmed by the design of sticky ends that determine which molecules associate, and how. The theory has been demonstrated experimentally using a system of synthetic DNA double-crossover molecules that self-assemble into two-dimensional crystals that have been visualized by atomic force microscopy. This experimental system provides an excellent platform for exploring the relationship between computation and molecular self-assembly, and thus represents a first step toward the ability to program molecular reactions and molecular structures

Construction, analysis, ligation, and self-assembly of DNA triple crossover complexes

This paper extends the study and prototyping of unusual DNA motifs, unknown in nature, but founded on principles derived from biological structures. Artificially designed DNA complexes show promise as building blocks for the construction of useful nanoscale structures, devices, and computers. The DNA triple crossover (TX) complex described here extends the set of experimentally characterized building blocks. It consists of four oligonucleotides hybridized to form three double-stranded DNA helices lying in a plane and linked by strand exchange at four immobile crossover points. The topology selected for this TX molecule allows for the presence of reporter strands along the molecular diagonal that can be used to relate the inputs and outputs of DNA-based computation. Nucleotide sequence design for the synthetic strands was assisted by the application of algorithms that minimize possible alternative base-pairing structures. Synthetic oligonucleotides were purified, stoichiometric mixtures were annealed by slow cooling, and the resulting DNA structures were analyzed by nondenaturing gel electrophoresis and heat-induced unfolding. Ferguson analysis and hydroxyl radical autofootprinting provide strong evidence for the assembly of the strands to the target TX structure. Ligation of reporter strands has been demonstrated with this motif, as well as the self-assembly of hydrogen-bonded two-dimensional crystals in two different arrangements. Future applications of TX units include the construction of larger structures from multiple TX units, and DNA-based computation. In addition to the presence of reporter strands, potential advantages of TX units over other DNA structures include space for gaps in molecular arrays, larger spatial displacements in nanodevices, and the incorporation of well-structured out-of-plane components in two-dimensional arrays

Towards Synthetic Life: Establishing a Minimal Segrosome for the Rational Design of Biomimetic Systems

DNA segregation is a fundamental life process, crucial for renewal, reproduction and propagation of all forms of life. Hence, a dedicated segregation machinery, a segrosome, must function reliably also in the context of a minimal cell. Conceptionally, the development of such a minimal cell follows a minimalistic approach, aiming at engineering a synthetic entity only consisting of the essential key elements necessary for a cell to survive. In this thesis, various prokaryotic segregation systems were explored as possible candidates for a minimal segrosome. Such a minimal segrosome could be applied for the rational design of biomimetic systems including, but not limited to, a minimal cell. DNA segregation systems of type I (ParABS) and type II (ParMRC) were compared for ensuring genetic stabilities in vivo using vectors derived from the natural secondary chromosome of Vibrio cholerae. The type II segregation system R1-ParMRC was chosen as the most promising candidate for a minimal segrosome, and it was characterized and reconstituted in vitro. This segregation system was encapsulated into biomimetic micro-compartments and its lifetime prolonged by coupling to ATP-regenerating as well as oxygen-scavenging systems. The segregation process was coupled to in vitro DNA replication using DNA nanoparticles as a mimic of the condensed state of chromosomes. Furthermore, another type II segregation system originating from the pLS20 plasmid from Bacillus subtilis (Alp7ARC) was reconstituted in vitro as a secondary orthogonal segrosome. Finally, a chimeric RNA segregation system was engineered that could be applied for an RNA-based protocell. Overall, this work demonstrates successful bottom-up assemblies of functional molecular machines that could find applications in biomimetic systems and lead to a deeper understanding of living systems