The (Un)Folding of Multidomain Proteins Through the Lens of Single-molecule Force-spectroscopy and Computer Simulation

Proteins are specialized molecules that catalyze most of the reactions that can sustain life, and they become functional by folding into a specific 3D structure. Despite their importance, the question, "how do proteins fold?" - first pondered in in the 1930's - is still listed as one of the top unanswered scientific questions as of 2005, according to the journal Science. Answering this question would provide a foundation for understanding protein function and would enable improved drug targeting, efficient biofuel production, and stronger biomaterials. Much of what we currently know about protein folding comes from studies on small, single-domain proteins, which may be quite different from the folding of large, multidomain proteins that predominate the proteomes of all organisms.

In this thesis I will discuss my work to fill this gap in understanding by studying the unfolding and refolding of large, multidomain proteins using the powerful combination of single-molecule force-spectroscopy experiments and molecular dynamic simulations.

The three model proteins studied - Luciferase, Protein S, and Streptavidin - lend insight into the inter-domain dependence for unfolding and the subdomain stabilization of binding ligands, and ultimately provide new insight into atomistic details of the intermediate states along the folding pathway.

Accuracy of computer-aided design/computer-assisted manufacture (CAD/CAM) fabricated dental restorations: a comparative study

Introduction: Computer-Aided-Design (CAD) and Computer-Aided-Manufacture (CAM) has been developed to fabricate fixed dental restorations accurately, faster and improve cost effectiveness of manufacture when compared to the conventional method. Two main methods exist in dental CAD/CAM technology: the subtractive and additive methods. While fitting accuracy of both methods has been explored, no study yet has compared the fabricated restoration (CAM output) to its CAD in terms of accuracy. The aim of this present study was to compare the output of various dental CAM routes to a sole initial CAD and establish the accuracy of fabrication. The internal fit of the various CAM routes were also investigated. The null hypotheses tested were: 1) no significant differences observed between the CAM output to the CAD and 2) no significant differences observed between the various CAM routes. Methods: An aluminium master model of a standard premolar preparation was scanned with a contact dental scanner (Incise, Renishaw, UK). A single CAD was created on the scanned master model (InciseCAD software, V2.5.0.140, UK). Twenty copings were then fabricated by sending the single CAD to a multitude of CAM routes. The copings were grouped (n=5) as: Laser sintered CoCrMo (LS), 5-axis milled CoCrMo (MCoCrMo), 3-axis milled zirconia (ZAx3) and 4-axis milled zirconia (ZAx4). All copings were micro-CT scanned (Phoenix X-Ray, Nanotom-S, Germany, power: 155kV, current: 60µA, 3600 projections) to produce 3-Dimensional (3D) models. A novel methodology was created to superimpose the micro-CT scans with the CAD (GOM Inspect software, V7.5SR2, Germany) to indicate inaccuracies in manufacturing. The accuracy in terms of coping volume was explored. The distances from the surfaces of the micro-CT 3D models to the surfaces of the CAD model (CAD Deviation) were investigated after creating surface colour deviation maps. Localised digital sections of the deviations (Occlusal, Axial and Cervical) and selected focussed areas were then quantitatively measured using software (GOM Inspect software, Germany). A novel methodology was also explored to digitally align (Rhino software, V5, USA) the micro-CT scans with the master model to investigate internal fit. Fifty digital cross sections of the aligned scans were created. Point-to-point distances were measured at 5 levels at each cross section. The five levels were: Vertical Marginal Fit (VF), Absolute Marginal Fit (AM), Axio-margin Fit (AMF), Axial Fit (AF) and Occlusal Fit (OF). Results: The results of the volume measurement were summarised as: VM-CoCrMo (62.8mm3 ) > VZax3 (59.4mm3 ) > VCAD (57mm3 ) > VZax4 (56.1mm3 ) > VLS (52.5mm3 ) and were all significantly different (p presented as areas with different colour. No significant differences were observed at the internal aspect of the cervical aspect between all groups of copings. Significant differences (p< M-CoCrMo Internal Occlusal, Internal Axial and External Axial 2 ZAx3 > ZAx4 External Occlusal, External Cervical 3 ZAx3 < ZAx4 Internal Occlusal 4 M-CoCrMo > ZAx4 Internal Occlusal and Internal Axial The mean values of AMF and AF were significantly (p M-CoCrMo and CAD > ZAx4. Only VF of M-CoCrMo was comparable with the CAD Internal Fit. All VF and AM values were within the clinically acceptable fit (120µm). Conclusion: The investigated CAM methods reproduced the CAD accurately at the internal cervical aspect of the copings. However, localised deviations at axial and occlusal aspects of the copings may suggest the need for modifications in these areas prior to fitting and veneering with porcelain. The CAM groups evaluated also showed different levels of Internal Fit thus rejecting the null hypotheses. The novel non-destructive methodologies for CAD/CAM accuracy and internal fit testing presented in this thesis may be a useful evaluation tool for similar applications.

3D Navigation with Six Degrees-of-Freedom using a Multi-Touch Display

With the introduction of new input devices, such as multi-touch surface displays, the Nintendo WiiMote, the Microsoft Kinect, and the Leap Motion sensor, among others, the field of Human-Computer Interaction (HCI) finds itself at an important crossroads that requires solving new challenges. Given the amount of three-dimensional (3D) data available today, 3D navigation plays an important role in 3D User Interfaces (3DUI). This dissertation deals with multi-touch, 3D navigation, and how users can explore 3D virtual worlds using a multi-touch, non-stereo, desktop display. The contributions of this dissertation include a feature-extraction algorithm for multi-touch displays (FETOUCH), a multi-touch and gyroscope interaction technique (GyroTouch), a theoretical model for multi-touch interaction using high-level Petri Nets (PeNTa), an algorithm to resolve ambiguities in the multi-touch gesture classification process (Yield), a proposed technique for navigational experiments (FaNS), a proposed gesture (Hold-and-Roll), and an experiment prototype for 3D navigation (3DNav). The verification experiment for 3DNav was conducted with 30 human-subjects of both genders. The experiment used the 3DNav prototype to present a pseudo-universe, where each user was required to find five objects using the multi-touch display and five objects using a game controller (GamePad). For the multi-touch display, 3DNav used a commercial library called GestureWorks in conjunction with Yield to resolve the ambiguity posed by the multiplicity of gestures reported by the initial classification. The experiment compared both devices. The task completion time with multi-touch was slightly shorter, but the difference was not statistically significant. The design of experiment also included an equation that determined the level of video game console expertise of the subjects, which was used to break down users into two groups: casual users and experienced users. The study found that experienced gamers performed significantly faster with the GamePad than casual users. When looking at the groups separately, casual gamers performed significantly better using the multi-touch display, compared to the GamePad. Additional results are found in this dissertation.

Fast 2D/3D object representation with growing neural gas

This work presents the design of a real-time system to model visual objects with the use of self-organising networks. The architecture of the system addresses multiple computer vision tasks such as image segmentation, optimal parameter estimation and object representation. We first develop a framework for building non-rigid shapes using the growth mechanism of the self-organising maps, and then we define an optimal number of nodes without overfitting or underfitting the network based on the knowledge obtained from information-theoretic considerations. We present experimental results for hands and faces, and we quantitatively evaluate the matching capabilities of the proposed method with the topographic product. The proposed method is easily extensible to 3D objects, as it offers similar features for efficient mesh reconstruction.

Application of computer-aided tomography techniques to visualize kelp holdfast structure reveals the importance of habitat complexity for supporting marine biodiversity

Ecosystem engineers that increase habitat complexity are keystone species in marine systems, increasing shelter and niche availability, and therefore biodiversity. For example, kelp holdfasts form intricate structures and host the largest number of organisms in kelp ecosystems. However, methods that quantify 3D habitat complexity have only seldom been used in marine habitats, and never in kelp holdfast communities. This study investigated the role of kelp holdfasts (Laminaria hyperborea) in supporting benthic faunal biodiversity. Computer-aided tomography (CT-) scanning was used to quantify the three-dimensional geometrical complexity of holdfasts, including volume, surface area and surface fractal dimension (FD). Additionally, the number of haptera, number of haptera per unit of volume, and age of kelps were estimated. These measurements were compared to faunal biodiversity and community structure, using partial least-squares regression and multivariate ordination. Holdfast volume explained most of the variance observed in biodiversity indices, however all other complexity measures also strongly contributed to the variance observed. Multivariate ordinations further revealed that surface area and haptera per unit of volume accounted for the patterns observed in faunal community structure. Using 3D image analysis, this study makes a strong contribution to elucidate quantitative mechanisms underlying the observed relationship between biodiversity and habitat complexity. Furthermore, the potential of CT-scanning as an ecological tool is demonstrated, and a methodology for its use in future similar studies is established. Such spatially resolved imager analysis could help identify structurally complex areas as biodiversity hotspots, and may support the prioritization of areas for conservation.

Application of computer-aided tomography techniques to visualize kelp holdfast structure reveals the importance of habitat complexity for supporting marine biodiversity

Ecosystem engineers that increase habitat complexity are keystone species in marine systems, increasing shelter and niche availability, and therefore biodiversity. For example, kelp holdfasts form intricate structures and host the largest number of organisms in kelp ecosystems. However, methods that quantify 3D habitat complexity have only seldom been used in marine habitats, and never in kelp holdfast communities. This study investigated the role of kelp holdfasts (Laminaria hyperborea) in supporting benthic faunal biodiversity. Computer-aided tomography (CT-) scanning was used to quantify the three-dimensional geometrical complexity of holdfasts, including volume, surface area and surface fractal dimension (FD). Additionally, the number of haptera, number of haptera per unit of volume, and age of kelps were estimated. These measurements were compared to faunal biodiversity and community structure, using partial least-squares regression and multivariate ordination. Holdfast volume explained most of the variance observed in biodiversity indices, however all other complexity measures also strongly contributed to the variance observed. Multivariate ordinations further revealed that surface area and haptera per unit of volume accounted for the patterns observed in faunal community structure. Using 3D image analysis, this study makes a strong contribution to elucidate quantitative mechanisms underlying the observed relationship between biodiversity and habitat complexity. Furthermore, the potential of CT-scanning as an ecological tool is demonstrated, and a methodology for its use in future similar studies is established. Such spatially resolved imager analysis could help identify structurally complex areas as biodiversity hotspots, and may support the prioritization of areas for conservation.

Face recognition using a unified 3D morphable model

We address the problem of 3D-assisted 2D face recognition in scenarios when the input image is subject to degradations or exhibits intra-personal variations not captured by the 3D model. The proposed solution involves a novel approach to learn a subspace spanned by perturbations caused by the missing modes of variation and image degradations, using 3D face data reconstructed from 2D images rather than 3D capture. This is accomplished by modelling the difference in the texture map of the 3D aligned input and reference images. A training set of these texture maps then defines a perturbation space which can be represented using PCA bases. Assuming that the image perturbation subspace is orthogonal to the 3D face model space, then these additive components can be recovered from an unseen input image, resulting in an improved fit of the 3D face model. The linearity of the model leads to efficient fitting. Experiments show that our method achieves very competitive face recognition performance on Multi-PIE and AR databases. We also present baseline face recognition results on a new data set exhibiting combined pose and illumination variations as well as occlusion.

Détection, reconstruction 3D et identification des êtres vivants du règne animal

L’automatisation de la détection et de l’identification des animaux est une tâche qui a de l’intérêt dans plusieurs domaines de recherche en biologie ainsi que dans le développement de systèmes de surveillance électronique. L’auteur présente un système de détection et d’identification basé sur la vision stéréo par ordinateur. Plusieurs critères sont utilisés pour identifier les animaux, mais l’accent a été mis sur l’analyse harmonique de la reconstruction en temps réel de la forme en 3D des animaux. Le résultat de l’analyse est comparé avec d’autres qui sont contenus dans une base évolutive de connaissances.

A Framework for Realistic 3D Tele-Immersion

Meeting, socializing and conversing online with a group of people using teleconferencing systems is still quite different from the experience of meeting face to face. We are abruptly aware that we are online and that the people we are engaging with are not in close proximity. Analogous to how talking on the telephone does not replicate the experience of talking in person. Several causes for these differences have been identified and we propose inspiring and innovative solutions to these hurdles in attempt to provide a more realistic, believable and engaging online conversational experience. We present the distributed and scalable framework REVERIE that provides a balanced mix of these solutions. Applications build on top of the REVERIE framework will be able to provide interactive, immersive, photo-realistic experiences to a multitude of users that for them will feel much more similar to having face to face meetings than the experience offered by conventional teleconferencing systems.

Analysis of Local Search Methods for 3D Data

In questa tesi sono stati analizzati alcuni metodi di ricerca per dati 3D. Viene illustrata una panoramica generale sul campo della Computer Vision, sullo stato dell’arte dei sensori per l’acquisizione e su alcuni dei formati utilizzati per la descrizione di dati 3D. In seguito è stato fatto un approfondimento sulla 3D Object Recognition dove, oltre ad essere descritto l’intero processo di matching tra Local Features, è stata fatta una focalizzazione sulla fase di detection dei punti salienti. In particolare è stato analizzato un Learned Keypoint detector, basato su tecniche di apprendimento di machine learning. Quest ultimo viene illustrato con l’implementazione di due algoritmi di ricerca di vicini: uno esauriente (K-d tree) e uno approssimato (Radial Search). Sono state riportate infine alcune valutazioni sperimentali in termini di efficienza e velocità del detector implementato con diversi metodi di ricerca, mostrando l’effettivo miglioramento di performance senza una considerabile perdita di accuratezza con la ricerca approssimata.

The Role of Art in Computer Game Design

Computer games are significant since they embody our youngsters’ engagement with contemporary culture, including both play and education. These games rely heavily on visuals, systems of sign and expression based on concepts and principles of Art and Architecture. We are researching a new genre of computer games, ‘Educational Immersive Environments’ (EIEs) to provide educational materials suitable for the school classroom. Close collaboration with subject teachers is necessary, but we feel a specific need to engage with the practicing artist, the art theoretician and historian. Our EIEs are loaded with multimedia (but especially visual) signs which act to direct the learner and provide the ‘game-play’ experience forming semiotic systems. We suggest the hypothesis that computer games are a space of deconstruction and reconstruction (DeRe): When players enter the game their physical world and their culture is torn apart; they move in a semiotic system which serves to reconstruct an alternate reality where disbelief is suspended. The semiotic system draws heavily on visuals which direct the players’ interactions and produce motivating gameplay. These can establish a reconstructed culture and emerging game narrative. We have recently tested our hypothesis and have used this in developing design principles for computer game designers. Yet there are outstanding issues concerning the nature of the visuals used in computer games, and so questions for contemporary artists. Currently, the computer game industry employs artists in a ‘classical’ role in production of concept sketches, storyboards and 3D content. But this is based on a specification from the client which restricts the artist in intellectual freedom. Our DeRe hypothesis places the artist at the generative centre, to inform the game designer how art may inform our DeRe semiotic spaces. This must of course begin with the artists’ understanding of DeRe in this time when our ‘identities are becoming increasingly fractured, networked, virtualized and distributed’ We hope to persuade artists to engage with the medium of computer game technology to explore these issues. In particular, we pose several questions to the artist: (i) How can particular ‘periods’ in art history be used to inform the design of computer games? (ii) How can specific artistic elements or devices be used to design ‘signs’ to guide the player through the game? (iii) How can visual material be integrated with other semiotic strata such as text and audio?

& she was like "O_O" : Animation of reported speech on Twitter

This study relates discourse-pragmatic aspects of the use of the quotatives say, be like, be all, and go to the question of the supposed or actual spoken-likeness of written computer-mediated communication (CMC). 1,800 tokens of reported speech, collected from Twitter, were analyzed in a “constructed dialogue” framework (Tannen, 2007). The results show that users of Twitter employ various CMC devices to animate and modally enrich reported speech, especially in speech reports with be like, be all, and go. They perform a style of communication that is reminiscent of conversational speech, even while having qualities that seem to belong uniquely to CMC.

Architectural-Physical Co-Design of 3D CPUs with Micro-Fluidic Cooling

The performance, energy efficiency and cost improvements due to traditional technology scaling have begun to slow down and present diminishing returns. Underlying reasons for this trend include fundamental physical limits of transistor scaling, the growing significance of quantum effects as transistors shrink, and a growing mismatch between transistors and interconnects regarding size, speed and power. Continued Moore's Law scaling will not come from technology scaling alone, and must involve improvements to design tools and development of new disruptive technologies such as 3D integration. 3D integration presents potential improvements to interconnect power and delay by translating the routing problem into a third dimension, and facilitates transistor density scaling independent of technology node. Furthermore, 3D IC technology opens up a new architectural design space of heterogeneously-integrated high-bandwidth CPUs. Vertical integration promises to provide the CPU architectures of the future by integrating high performance processors with on-chip high-bandwidth memory systems and highly connected network-on-chip structures. Such techniques can overcome the well-known CPU performance bottlenecks referred to as memory and communication wall. However the promising improvements to performance and energy efficiency offered by 3D CPUs does not come without cost, both in the financial investments to develop the technology, and the increased complexity of design. Two main limitations to 3D IC technology have been heat removal and TSV reliability. Transistor stacking creates increases in power density, current density and thermal resistance in air cooled packages. Furthermore the technology introduces vertical through silicon vias (TSVs) that create new points of failure in the chip and require development of new BEOL technologies. Although these issues can be controlled to some extent using thermal-reliability aware physical and architectural 3D design techniques, high performance embedded cooling schemes, such as micro-fluidic (MF) cooling, are fundamentally necessary to unlock the true potential of 3D ICs. A new paradigm is being put forth which integrates the computational, electrical, physical, thermal and reliability views of a system. The unification of these diverse aspects of integrated circuits is called Co-Design. Independent design and optimization of each aspect leads to sub-optimal designs due to a lack of understanding of cross-domain interactions and their impacts on the feasibility region of the architectural design space. Co-Design enables optimization across layers with a multi-domain view and thus unlocks new high-performance and energy efficient configurations. Although the co-design paradigm is becoming increasingly necessary in all fields of IC design, it is even more critical in 3D ICs where, as we show, the inter-layer coupling and higher degree of connectivity between components exacerbates the interdependence between architectural parameters, physical design parameters and the multitude of metrics of interest to the designer (i.e. power, performance, temperature and reliability). In this dissertation we present a framework for multi-domain co-simulation and co-optimization of 3D CPU architectures with both air and MF cooling solutions. Finally we propose an approach for design space exploration and modeling within the new Co-Design paradigm, and discuss the possible avenues for improvement of this work in the future.

Physical Design Methodologies for Low Power and Reliable 3D ICs

As the semiconductor industry struggles to maintain its momentum down the path following the Moore's Law, three dimensional integrated circuit (3D IC) technology has emerged as a promising solution to achieve higher integration density, better performance, and lower power consumption. However, despite its significant improvement in electrical performance, 3D IC presents several serious physical design challenges. In this dissertation, we investigate physical design methodologies for 3D ICs with primary focus on two areas: low power 3D clock tree design, and reliability degradation modeling and management. Clock trees are essential parts for digital system which dissipate a large amount of power due to high capacitive loads. The majority of existing 3D clock tree designs focus on minimizing the total wire length, which produces sub-optimal results for power optimization. In this dissertation, we formulate a 3D clock tree design flow which directly optimizes for clock power. Besides, we also investigate the design methodology for clock gating a 3D clock tree, which uses shutdown gates to selectively turn off unnecessary clock activities. Different from the common assumption in 2D ICs that shutdown gates are cheap thus can be applied at every clock node, shutdown gates in 3D ICs introduce additional control TSVs, which compete with clock TSVs for placement resources. We explore the design methodologies to produce the optimal allocation and placement for clock and control TSVs so that the clock power is minimized. We show that the proposed synthesis flow saves significant clock power while accounting for available TSV placement area. Vertical integration also brings new reliability challenges including TSV's electromigration (EM) and several other reliability loss mechanisms caused by TSV-induced stress. These reliability loss models involve complex inter-dependencies between electrical and thermal conditions, which have not been investigated in the past. In this dissertation we set up an electrical/thermal/reliability co-simulation framework to capture the transient of reliability loss in 3D ICs. We further derive and validate an analytical reliability objective function that can be integrated into the 3D placement design flow. The reliability aware placement scheme enables co-design and co-optimization of both the electrical and reliability property, thus improves both the circuit's performance and its lifetime. Our electrical/reliability co-design scheme avoids unnecessary design cycles or application of ad-hoc fixes that lead to sub-optimal performance. Vertical integration also enables stacking DRAM on top of CPU, providing high bandwidth and short latency. However, non-uniform voltage fluctuation and local thermal hotspot in CPU layers are coupled into DRAM layers, causing a non-uniform bit-cell leakage (thereby bit flip) distribution. We propose a performance-power-resilience simulation framework to capture DRAM soft error in 3D multi-core CPU systems. In addition, a dynamic resilience management (DRM) scheme is investigated, which adaptively tunes CPU's operating points to adjust DRAM's voltage noise and thermal condition during runtime. The DRM uses dynamic frequency scaling to achieve a resilience borrow-in strategy, which effectively enhances DRAM's resilience without sacrificing performance. The proposed physical design methodologies should act as important building blocks for 3D ICs and push 3D ICs toward mainstream acceptance in the near future.

Contributions to 3D Data Registration and Representation

Nowadays, new computers generation provides a high performance that enables to build computationally expensive computer vision applications applied to mobile robotics. Building a map of the environment is a common task of a robot and is an essential part to allow the robots to move through these environments. Traditionally, mobile robots used a combination of several sensors from different technologies. Lasers, sonars and contact sensors have been typically used in any mobile robotic architecture, however color cameras are an important sensor due to we want the robots to use the same information that humans to sense and move through the different environments. Color cameras are cheap and flexible but a lot of work need to be done to give robots enough visual understanding of the scenes. Computer vision algorithms are computational complex problems but nowadays robots have access to different and powerful architectures that can be used for mobile robotics purposes. The advent of low-cost RGB-D sensors like Microsoft Kinect which provide 3D colored point clouds at high frame rates made the computer vision even more relevant in the mobile robotics field. The combination of visual and 3D data allows the systems to use both computer vision and 3D processing and therefore to be aware of more details of the surrounding environment. The research described in this thesis was motivated by the need of scene mapping. Being aware of the surrounding environment is a key feature in many mobile robotics applications from simple robotic navigation to complex surveillance applications. In addition, the acquisition of a 3D model of the scenes is useful in many areas as video games scene modeling where well-known places are reconstructed and added to game systems or advertising where once you get the 3D model of one room the system can add furniture pieces using augmented reality techniques. In this thesis we perform an experimental study of the state-of-the-art registration methods to find which one fits better to our scene mapping purposes. Different methods are tested and analyzed on different scene distributions of visual and geometry appearance. In addition, this thesis proposes two methods for 3d data compression and representation of 3D maps. Our 3D representation proposal is based on the use of Growing Neural Gas (GNG) method. This Self-Organizing Maps (SOMs) has been successfully used for clustering, pattern recognition and topology representation of various kind of data. Until now, Self-Organizing Maps have been primarily computed offline and their application in 3D data has mainly focused on free noise models without considering time constraints. Self-organising neural models have the ability to provide a good representation of the input space. In particular, the Growing Neural Gas (GNG) is a suitable model because of its flexibility, rapid adaptation and excellent quality of representation. However, this type of learning is time consuming, specially for high-dimensional input data. Since real applications often work under time constraints, it is necessary to adapt the learning process in order to complete it in a predefined time. This thesis proposes a hardware implementation leveraging the computing power of modern GPUs which takes advantage of a new paradigm coined as General-Purpose Computing on Graphics Processing Units (GPGPU). Our proposed geometrical 3D compression method seeks to reduce the 3D information using plane detection as basic structure to compress the data. This is due to our target environments are man-made and therefore there are a lot of points that belong to a plane surface. Our proposed method is able to get good compression results in those man-made scenarios. The detected and compressed planes can be also used in other applications as surface reconstruction or plane-based registration algorithms. Finally, we have also demonstrated the goodness of the GPU technologies getting a high performance implementation of a CAD/CAM common technique called Virtual Digitizing.