Abstract Models of Molecular Walkers

Recent advances in single-molecule chemistry have led to designs for artificial multi-pedal walkers that follow tracks of chemicals. The walkers, called molecular spiders, consist of a rigid chemically inert body and several flexible enzymatic legs. The legs can reversibly bind to chemical substrates on a surface, and through their enzymatic action convert them to products. We study abstract models of molecular spiders to evaluate how efficiently they can perform two tasks: molecular transport of cargo over tracks and search for targets on finite surfaces. For the single-spider model our simulations show a transient behavior wherein certain spiders move superdiffusively over significant distances and times. This gives the spiders potential as a faster-than-diffusion transport mechanism. However, analysis shows that single-spider motion eventually decays into an ordinary diffusive motion, owing to the ever increasing size of the region of products. Inspired by cooperative behavior of natural molecular walkers, we propose a symmetric exclusion process (SEP) model for multiple walkers interacting as they move over a one-dimensional lattice. We show that when walkers are sequentially released from the origin, the collective effect is to prevent the leading walkers from moving too far backwards. Hence, there is an effective outward pressure on the leading walkers that keeps them moving superdiffusively for longer times. Despite this improvement the leading spider eventually slows down and moves diffusively, similarly to a single spider. The slowdown happens because all spiders behind the leading spiders never encounter substrates, and thus they are never biased. They cannot keep up with leading spiders, and cannot put enough pressure on them. Next, we investigate search properties of a single and multiple spiders moving over one- and two-dimensional surfaces with various absorbing and reflecting boundaries. For the single-spider model we evaluate by how much the slowdown on newly visited sites, owing to catalysis, can improve the mean first passage time of spiders and show that in one dimension, when both ends of the track are an absorbing boundary, the performance gain is lower than in two dimensions, when the absorbing boundary is a circle; this persists even when the absorbing boundary is a single site. Next, we study how multiple molecular spiders influence one another during the search. We show that when one spider reaches the trace of another spider it is more likely not to follow the trace and instead explore unvisited sites. This interaction between the spiders gives them an advantage over independent random walkers in a search for multiple targets. We also study how efficiently the spiders with various gaits are able to find specific targets. Spiders with gaits that allow more freedom of leg movement find their targets faster than spiders with more restrictive gaits. For every gait, there is an optimal detachment rate that minimizes the time to find all target sites

Logic Circuits Based on Extended Molecular Spider Systems

Spatial locality brings the advantages of computation speed-up and sequence reuse to molecular computing. In particular, molecular walkers that undergo localized re- actions are of interest for implementing logic computations at the nanoscale. We use molecular spider walkers to implement logic circuits. We develop an extended multi- spider model with a dynamic environment wherein signal transmission is triggered via localized reactions, and use this model to implement three basic gates (AND, OR, and NOT) and a cascading mechanism. We develop an algorithm to automatically generate the layout of the circuit. We use a kinetic Monte Carlo algorithm to simulate circuit computations, and we analyze circuit complexity: our design scales linearly with formula size and has a logarithmic time complexity

The Hierarchical Structure, Dynamics and Assembly of Spider Silks

Spider silks are biological protein polymers which are spun into fibers with an incredibly diverse range of mechanical properties and functions. Spider silks have long been recognized to have mechanical properties that rival most, if not all man-made materials. However, it is not yet possible to make synthetic spider silk fibers that mimic the impressive properties of their native counterparts. This is due in part to the hierarchical nature of spider silk, which introduces a complex organization of structures at nearly every length scale from the atomic to the micron level. The research in this dissertation has been in pursuit of understanding the structure, dynamics and assembly of silk proteins into high performance fibers at the molecular and nanoscale levels. The mechanical properties of a number of spider silks were measured and are discussed and placed intheir biological context. The underlying molecular interactions that give rise to these fibers are investigated by DLS, cryo-TEM, solution and solid-state NMR. The solution NMR work combined with cryo-TEM illustrates that silk proteins are stored as protein pre-assemblies in the silk gland and are the fundamental precursors to silk fibrils. The application of denaturant was used to disrupt these protein superstructures and understand the forces and dynamics that facilitate assembly. Finally, solid-state NMR was used to structurally characterize prey wrap spider silk, the toughest of the spider silks, and illustrate its α-helical coiled-coil hierarchical structure that helps explain this silk’s high extensibility and impressive mechanical performance

Design Of Dna Strand Displacement Based Circuits

DNA is the basic building block of any living organism. DNA is considered a popular candidate for future biological devices and circuits for solving genetic disorders and several other medical problems. With this objective in mind, this research aims at developing novel approaches for the design of DNA based circuits. There are many recent developments in the medical field such as the development of biological nanorobots, SMART drugs, and CRISPR-Cas9 technologies. There is a strong need for circuits that can work with these technologies and devices. DNA is considered a suitable candidate for designing such circuits because of the programmability of the DNA strands, small size, lightweight, known thermodynamics, higher parallelism, and exponentially reducing the cost of synthesizing techniques. The DNA strand displacement operation is useful in developing circuits with DNA strands. The circuit can be either a digital circuit, in which the logic high and logic low states of the DNA strand concentrations are considered as the signal, or it can be an analog circuit in which the concentration of the DNA strands itself will act as the signal. We developed novel approaches in this research for the design of digital, as well as analog circuits keeping in view of the number of DNA strands required for the circuit design. Towards this goal in the digital domain, we developed spatially localized DNA majority logic gates and an inverter logic gate that can be used with the existing seesaw based logic gates. The majority logic gates proposed in this research can considerably reduce the number of strands required in the design. The introduction of the logic inverter operation can translate the dual rail circuit architecture into a monorail architecture for the seesaw based logic circuits. It can also reduce the number of unique strands required for the design into approximately half. The reduction in the number of unique strands will consequently reduce the leakage reactions, circuit complexity, and cost associated with the DNA circuits. The real world biological inputs are analog in nature. If we can use those analog signals directly in the circuits, it can considerably reduce the resources required. Even though analog circuits are highly prone to noise, they are a perfect candidate for performing computations in the resource-limited environments, such as inside the cell. In the analog domain, we are developing a novel fuzzy inference engine using analog circuits such as the minimum gate, maximum gate, and fan-out gates. All the circuits discussed in this research were designed and tested in the Visual DSD software. The biological inputs are inherently fuzzy in nature, hence a fuzzy based system can play a vital role in future decision-making circuits. We hope that our research will be the first step towards realizing these larger goals. The ultimate aim of our research is to develop novel approaches for the design of circuits which can be used with the future biological devices to tackle many medical problems such as genetic disorders

(Non)Parallel Evolution

Parallel evolution across replicate populations has provided evolutionary biologists with iconic examples of adaptation. When multiple populations colonize seemingly similar habitats, they may evolve similar genes, traits, or functions. Yet, replicated evolution in nature or in the laboratory often yields inconsistent outcomes: Some replicate populations evolve along highly similar trajectories, whereas other replicate populations evolve to different extents or in distinct directions. To understand these heterogeneous outcomes, biologists are increasingly treating parallel evolution not as a binary phenomenon but rather as a quantitative continuum ranging from parallel to nonparallel. By measuring replicate populations’ positions along this (non)parallel continuum, we can test hypotheses about evolutionary and ecological factors that influence the extent of repeatable evolution. We review evidence regarding the manifestation of (non)parallel evolution in the laboratory, in natural populations, and in applied contexts such as cancer. We enumerate the many genetic, ecological, and evolutionary processes that contribute to variation in the extent of parallel evolution

Venom Yield, Regeneration, and Composition in the Centipede Scolopendra Polymorpha

In this dissertation, I investigated yield, regeneration, and composition of centipede venom. In the first of three empirical studies, I investigated how size influenced venom volume yield and protein concentration in Scolopendra polymorpha and S. subspinipes. I also examined additional potential influences on yield in S. polymorpha, including relative forcipule size, relative mass, geographic origin, sex, time in captivity, and milking history. Volume yield was positively linearly related to body length in both species; however, body length and protein concentration were uncorrelated. In S. polymorpha, yield was most influenced by body length, but was also positively associated with relative forcipule length and relative body mass. In the second study, I investigated venom volume and total protein regeneration during the 14-day period subsequent to venom extraction in S. polymorpha. I further tested the hypothesis that venom protein components, separated by RP-FPLC, undergo asynchronous synthesis. During the first 48 hours, volume and protein mass increased linearly. However, protein regeneration lagged behind volume regeneration, with only 65–86% of venom volume and 29–47% of protein mass regenerated during the first 2 days. No significant additional regeneration occurred over the subsequent 12 days. Analysis of chromatograms of individual venom samples revealed that five of 10 chromatographic regions and 12 of 28 peaks demonstrated changes in percent of total peak area among milking intervals, indicating that venom proteins are regenerated asynchronously. In the third study, I characterized the venom composition of S. polymorpha using proteomic methods. I demonstrated that the venom of S. polymorpha is complex, generating 23 bands by SDS-PAGE and 56 peaks by RP-FPLC. MALDI TOF MS revealed hundreds of components with masses ranging from 1014.5 to 82863.9 Da. The distribution of molecular masses was skewed toward smaller peptides and proteins, with 72% of components found below 12 kDa. BLASTp sequence similarity searching of MS/MSderived amino acid sequences demonstrated 20 different sequences with similarity to known venom components, including serine proteases, ion-channel activators/inhibitors, and neurotoxins. In Appendix A, I reviewed how animals strategically deploy various emissions, including venom, highlighting how the metabolic and ecological value of these emissions leads to their judicious use

Insect-selective spider toxins targeting voltage-gated sodium channels

The voltage-gated sodium (Nav) channel is a target for a number of drugs, insecticides and neurotoxins. These bind to at least seven identified neurotoxin binding sites and either block conductance or modulate Nav channel gating. A number of peptide neurotoxins from the venoms of araneomorph and mygalomorph spiders have been isolated and characterized and determined to interact with several of these sites. These all conform to an 'inhibitor cystine-knot' motif with structural, but not sequence homology, to a variety of other spider and marine snail toxins. Of these, spider toxins several show phyla-specificity and are being considered as lead compounds for the development of biopesticides. Hainantoxin-I appears to target site-1 to block Nav channel conductance. Magi 2 and Tx4(6-1) slow Nav channel inactivation via an interaction with site-3. The δ-palutoxins, and most likely μ-agatoxins and curtatoxins, target site-4. However, their action is complex with the μ-agatoxins causing a hyperpolarizing shift in the voltage-dependence of activation, an action analogous to scorpion β-toxins, but with both δ-palutoxins and μ-agatoxins slowing Nav channel inactivation, a site-3-like action. In addition, several other spider neurotoxins, such as δ-atracotoxins, are known to target both insect and vertebrate Nav channels most likely as a result of the conserved structures within domains of voltage-gated ion channels across phyla. These toxins may provide tools to establish the molecular determinants of invertebrate selectivity. These studies are being greatly assisted by the determination of the pharmacophore of these toxins, but without precise identification of their binding site and mode of action their potential in the above areas remains underdeveloped. © 2006 Elsevier Ltd. All rights reserved