Foram determinadas por meio de difração de raios x as estruturas de cinco compostos azaçúcares. Foram estudadas as interações envolvidas na formação das redes cristalinas em cada um dos compostos analisados. Foi encontrado que nos compostos azaçúcares estudados, as interações principais são as ligações de hidrogênio do tipo C-H···O e C-H···π. Este comportamento foi verificado usando ferramentas como as superfícies de Hirshfeld e os gráficos de impressão digital. Realizou-se o estudo de docking molecular dos compostos azaçúcares com respeito à enzima purina nucleosídeo fosforilase (PNP). Foi determinado que estes compostos têm a capacidade de entrar no sitio ativo da PNP. O estudo das interações dos cinco azaçúcares com a PNP mostrou que estes compostos apresentam as mesmas interações presentes em inibidores da PNP já reportados. / Structures of five azasugars were determined by X-ray diffraction. Crystal network interactions were analyzed for each compound. The main interaction found for these azasugar compounds is hydrogen bond as C-H···O e C-H···π. This behavior was verified by tools as Hirshfeld surface and 2D finger print plots. Molecular docking was performed for azasugar compounds in Purine Nucleoside phosphorylase (PNP). This study confirmed that these compounds are available to enter to the PNP active site. Interactions exploration showed the same interactions for the azasugars studied and for already known PNP inhibitors.

Protein-protein and protein-ligand interactions are fundamental as many proteins mediate their biological function through these interactions. Many important applications follow directly from the identification of residues in the interfaces between protein-protein and protein-ligand interactions, such as drug design, protein mimetic engineering, elucidation of molecular pathways, and understanding of disease mechanisms. The identification of interface residues can also guide the docking process to build the structural model of protein-protein complexes. This dissertation focuses on developing computational approaches for protein-ligand and protein-protein binding site prediction and applying these predictions to improve protein-protein docking. First, we develop an automated approach LIGSITEcs to predict protein-ligand binding site, based on the notion of surface-solvent-surface events and the degree of conservation of the involved surface residues. We compare our algorithm to four other approaches, LIGSITE, CAST, PASS, and SURFNET, and evaluate all on a dataset of 48 unbound/bound structures and 210 bound-structures. LIGSITEcs performs slightly better than the other tools and achieves a success rate of 71% and 75%, respectively. Second, for protein-protein binding site, we develop metaPPI, a meta server for interface prediction. MetaPPI combines results from a number of tools, such as PPI_Pred, PPISP, PINUP, Promate, and SPPIDER, which predict enzyme-inhibitor interfaces with success rates of 23% to 55% and other interfaces with 10% to 28% on a benchmark dataset of 62 complexes. After refinement, metaPPI significantly improves prediction success rates to 70% for enzyme-inhibitor and 44% for other interfaces. Third, for protein-protein docking, we develop a FFT-based docking algorithm and system BDOCK, which includes specific scoring functions for specific types of complexes. BDOCK uses family-based residue interface propensities as a scoring function and obtains improvement factors of 4-30 for enzyme-inhibitor and 4-11 for antibody-antigen complexes in two specific SCOP families. Furthermore, the degrees of buriedness of surface residues are integrated into BDOCK, which improves the shape discriminator for enzyme-inhibitor complexes. The predicted interfaces from metaPPI are integrated as well, either during docking or after docking. The evaluation results show that reliable interface predictions improve the discrimination between near-native solutions and false positive. Finally, we propose an implicit method to deal with the flexibility of proteins by softening the surface, to improve docking for non enzyme-inhibitor complexes.

Dissertation (PhD)--Stellenbosch, 2004. / ENGLISH ABSTRACT: The dissertation reflects the research done on the design principles and success factors for the operation of cross dock facilities in grocery and retail supply chains. The cross dock is a particular facility in the supply chain where goods are received from suppliers, sorted without storage of the goods, and then efficiently moved to downstream customers. Cross docks are not a new operation. However, the use in high volume grocery and retail operational capabilities is poorly understood and is not uniquely defined. The problem is that cross docks are often seen as extensions of warehouses. The same personnel, systems and processes are applied and the efficiency potential of the cross dock is not achieved. Warehouses are orientated towards storing the full range of product and allowing the pick to be done from this storage buffer to provide any or all of these products to a customer. Cross docks will only handle products that are used in larger quantities and that are sent to most, if not all, the customers. The cross dock is therefore distinct and very different from the traditional warehouse. The published research tends to focus on the technical aspects of the cross dock layout. This research is primarily in the scheduling of the trucks into the yard of the facility; the allocation of trucks to specific doors of the facility; and the allocation of doors to receiving and despatch functions within the facility. Very little information or research reflects the design principles and success factors for the cross dock and its supply chain. The only classification of the cross dock in the literature is whether the barcode is added to the item before or after receipt at the cross dock. For this research work a literature survey was conducted and five major operations were reviewed, in South Africa and the USA. The research empirically drew logical conclusions, which were tested in the operations and found to be correct. This allowed the design principles and success factors to be determined for a successful cross dock. The research extends the knowledge of the cross dock operation and design: - • A new classification for the feasible types of cross docks in the supply chain was developed. Three factors are shown to be of primary importance: - o Where in the supply chain the identification of specific items for a customer is done; o Where the sort is done for the items to be delivered to a customer; and o Whether the supplier is providing one product or multiple products to the sort. From these three factors, eight potential classifications could be defined. However, only three practical types of cross dock can be determined from these eight alternatives. These are named in this research as Cross Dock Managed Load (CML); Joint Managed Load (JML); and the Supplier Managed Load (SML). The cross dock is far more effective than the warehouse when the total work (excluding inventory) is considered. The earlier in the supply chain the product is identified for the use of the entire downstream supply chain, the more effective will be the total supply chain. Thus the greatest supply chain effectiveness possible is with the SML, then the JML and finally the CML. • The operation of a cross dock is very similar to a continuous manufacturing process. There is no buffer of stock to decouple the inbound and outbound processes, and the operation takes place in a restricted area. However, in the retail chain, the workload alters with different orders and different days. Daily load differences vary by as much as 90%. This results in vastly different workloads and variations of throughput. This is similar to a batch operation with highly variable workloads between batches. The literature recommends the use of Just in Time (JIT) practice for cross docks. This is inappropriate as its primary requirements are continuous full volume operation and continuous small improvements to achieve a balanced operation. The most appropriate method of process improvement is the Theory of Constraints (TO C) and not JlT. • The management must have a detailed, disciplined approach. This implies standardised methods of operation, and a high degree of training. Equally there is the requirement for a special type of personnel to operate the cross dock. These operating personnel must be able to operate with precision (i.e. very low error rates) and be able to maintain this capability for continuous periods. • The systems required for a successful operation must include the capabilities of Yard Management, WMS for cross docking, Order Management with Advanced Shipping Notice (ASN) capability and Track and Trace across the supply chain. The items need to be identified by a barcode. The information required on the barcode will be determined by. the information systems capability of the least advanced service provider in the supply chain. If this service provider can receive and transmit all the data required for the supply chain from and to the other members, then the barcode need only be an identification number of the specific item. The data pertaining to the items is then passed from system to system in the supply chain. If data movement is not possible between all the parties in the entire supply chain, then the barcode must contain the information that will identify the item, the origin and the final delivery destination. If the items are delivered as part of a consignment, a further quantum of information is required to identify the total number of items in the consignment and the specific item within the consignment. • The research shows that the overall capability of the cross dock or its maximum capacity is the combination of the capability of the personnel and the cross dock design. Restrictions on either the personnel capability or the design of the cross dock, or both, severely reduces the effectiveness of the cross dock. • The previous research on the sequence of allocation of trucks to specific doors within the cross dock can be enhanced with a new sequencing method. The new method allocates the transport, in sequence of arrival, to the open door that either numrruses the walk distance in the facility; or maximises the completion of the consignments in order to minimise the area required to build the consignments; or a combination of both. The choice of these will be determined by the constraints imposed by the design of the building. This is an important extension as this ties the supply chain into the cross dock operation, rather than looking at the cross dock in isolation as has been done in this previous research. • The factors that influence the design of a cross dock as to its size, shape, number of doors, and the specifically required additional areas, is defined in detail. The principles of these factors and their inter-relationships and dependencies are used in a detailed design for a cross dock. The detailed design process is set out from data analysis through to the actual size calculations and layouts. Measurements of walk distance and sort movement are used to determine the most effective design. The design is shown to be considerably more effective than the older designs. This work has significantly extended the research on the design principles and success factors for implementation of cross docks in retail supply chains. The research derives a unique new classification for cross docks. An improvement is made to existing research on the allocation of the transport to particular doors in the cross dock. The operation, management and personnel are shown to require specific characteristics. The information systems required for effective cross docks is determined and defined. The identification of the individual items by barcode and the information required within the barcode depending on the information sophistication of the service providers in the supply chain is defined. A detail process to design a cross dock is evolved, with the full knowledge of the factors that must be considered and their interrelationships. Measurements to determine the effectiveness of the design are used to choose the most appropriate design. All these are then synthesised into a new design, which is far more effective than any of the other designs researched. The design process will produce a very effective cross dock as has been demonstrated with a new facility. / AFRIKAANSE OPSOMMING: geen opsomming

Thesis (MEng (Electrical and Electronic Engineering))--University of Stellenbosch, 2009. / Autonomous rendezvouz and docking is seen as an enabling technology. It allows, among others, the construction of larger space platforms in-orbit and also provides a means for the in-orbit servicing of space vehicles. In this thesis a docking sequence is proposed and tested in both simulation and practice. This therefore also requires the design and construction of a test platform. A model hovercraft is used to emulate the chaser satellite in a 2-dimensional plane as it moves relatively frictionlessly. The hovercraft is also equipped with a single camera (monocular vision) that is used as the main sensor to estimate the target’s pose (relative position and orientation). An imitation of a target satellite was made and equipped with light markers that are used by the chaser’s camera sensor. The position of the target’s lights in the image is used to determine the target’s pose using a modified version ofMalan’s Extended Kalman Filter [20]. This information is then used during the docking sequence. This thesis successfully demonstrated the autonomous and reliable identification of the target’s lights in the image, and the autonomous docking of a satellite pair using monocular camera vision in both simulation and emulation.

The overall aim of this work was to apply HINT, an empirical scoring function based on the understanding of hydrophobicity, to analyze and predict the binding affinities and biological activities of colchicine-site anticancer agents. The second, concurrent aim was to improve the scoring function by incorporating tautomerism within the modeling process. Our belief is that proper evaluation of tautomeric forms for small molecules will improve performance of virtual screening. The novel pyrrole-based compounds targeting the colchicine site were docked into the receptor using HINT as a rescoring function. Two distinct binding modes dictated by the size and shape of a subpocket were predicted to differentiate the highly active compounds from the weak ones. Of the residues predicted to participate in binding for the active binding mode, Cys241β was revealed to form a weak but critical hydrogen bond with the ligand. A larger collection of colchicine-site agents, biologically tested in the same laboratory including our pyrrole-based compounds were subject to 3D quantitative structure-activity relationship (QSAR) study. Using results on docking the pyrrole compounds as a guide, relative binding poses and QSAR models were built to facilitate ligand design and optimization. A new 3D modeling approach was introduced to visually highlight the unique features of highly active compounds and the commonality of all compounds in the dataset using HINT maps and successfully tested on the colchicine-site agents. These results will provide valuable guidance in the future design and development of new colchicine-site agents. To incorporate tautomerism within HINT, we proposed and developed two workflow approaches: a general search tool using a simple and intuitive algorithm analyzing hydrogen shift patterns to identify and enumerate tautomeric structures, and a database that contains commonly observed tautomeric structures. The first approach was designed for small-scale docking studies and the second approach was designed for large-scale virtual screening. The tautomer module in HINT will give more accurate modeling results when the compound encountered is able to tautomerize.

Thèse numérisée par la Direction des bibliothèques de l'Université de Montréal.

Les travaux présentés dans ce mémoire se situent dans le cadre général de la recherche de nouveaux médicaments par le biais de techniques informatiques. La première partie de ce document est centrée autour du développement de la plateforme logicielle VSM-G (Virtual Screening Manager for Grids). Le but poursuivi par ce projet est de fournir un outil convivial et simple d'utilisation afin de conduire des études de criblage virtuel à haut-débit. Le coeur de VSM-G repose sur une stratégie multi-étapes de filtres successifs permettant le traitement efficace de chimiothèques de grande taille. Deux filtres ont été utilisés pour ce travail et implémentés dans VSM-G : un programme innovant d’estimation rapide de complémentarité géométrique entre molécules-candidates et site actif (SHEF) précéde un algorithme de docking flexible plus conventionnel (GOLD). Les avantages de cette méthodologie, associée à la prise en charge de multiples conformations de la cible étudiée (le récepteur nucléaire LXRß), sont présentés tout d’abord par une étude de preuve de concept, puis à travers une campagne de criblage virtuel à grande échelle. L'autre partie de ces travaux, exclusivement applicative, concerne l'étude du domaine FAT de la kinase d'adhérence focale FAK. FAK est une cible d’intérêt pharmaceutique particulièrement intéressante, car clairement impliquée dans divers processus de développement cancéreux. Le but de cette étude est double : il s’agit tout d’abord de mieux comprendre le mode de fonctionnement du domaine FAT de FAK à travers une étude biophysique pour en évaluer la flexibilité ; et ensuite concevoir in silico des petites molécules peptidomimétiques permettant de moduler son activité, ce qui pourrait limiter une progression tumorale. / The work presented here deals with drug discovery by means of computational techniques. The first part is focused around the development of the VSM-G (Virtual Screening Manager for Grids) software platform. This project aims to provide a user-friendly and easy-to-use tool for performing high throughput virtual screening experiments. The core of VSM-G is a multiple-step screening strategy in which several filters are organized sequentially as to tackle large chemical libraries efficiently. Two filters were used for this study and implemented into VSM-G: a new and fast ligand-active site geometrical complementarity estimation program (SHEF) precedes a conventional flexible docking tool (GOLD). We describe the advantages of such an approach, associated with the use of multiple target conformations for the LXRß nuclear receptor, by presenting a proof-of-concept study. A high-throughput virtual screening campaign is then performed. The second part of this work, exclusively applicative, deals with the study of the FAT domain of the focal adhesion kinase (FAK). FAK is an important pharmaceutical target due to its involvement in the development of various forms of cancer. The first goal is to gain knowledge regarding FAT flexibility and active state structural properties. The second objective is to design in silico peptidomimetic compounds targeting FAT and therefore potentially modulate FAK activity during tumour progression.

Thyroid disruptions by xenobiotics have been associated with a broad spectrum of severe adverse human health effects, such as impaired brain development and metabolic syndrome. Ingestion of indoor dust and contact with industrial chemicals are two significant human exposure routes of thyroid hormone disrupting chemicals (THDCs), raising serious concerns for human health. However, it is a laborious and costly process to identify THDCs using conventional experimental methods, due to the number of chemicals in commerce and the varieties of potential disruption mechanisms. In this thesis, we are aimed at in silico identification of novel THDCs targeting transthyretin (TTR) and thyroid hormone receptor (THR) among dust contaminants and commonly used industrial chemicals. In vitro assays were used to validate the in silico prediction results. Co-crystallization and molecular dynamics (MD) simulations were applied to reveal binding modes of THDCs at the studied biological targets and to explain their intermolecular recognition. The main findings presented in this thesis are: 1. Over 144 environmental pollutants have been confirmed as TTR-binders in vitro and these cover a wide range of environmental pollutants and show distinct chemical profiles including a large group of halogenated aromatic compounds and a second group of per- and polyfluoroalkyl substances. (Paper I) 2. In total 485 organic contaminants have been reported to be detected in household dust. The developed QSAR classification model predicted 7.6% of these dust contaminants and 53.1% of their metabolites as potential TTR-binders, which emphasizes the importance of metabolic bioactivation. After in vitro validation, four novel TTR binders with IC50 ≤ 10 µM were identified, i.e. perfluoroheptanesulfonic acid, 2,4,2',4'-tetrahydroxybenzophenone (BP2), 2,4,5-trichlorophenoxyacetic acid, and 3,5,6-trichloro-2-pyridinol. (Paper II) 3. The development of a robust structure-based virtual screening (VS) protocol resulted in the prediction of 31 dust contaminants as potential binders to THRβ1 including musk compounds, PFASs, and bisphenol A derivatives. The in vitro experiments confirmed four compounds as weak binders to THRβ1, i.e. 2,4,5-trichlorophenoxyacetic acid, bisphenol A (3-chloro-2-hydroxypropyl) (2,3-dihydroxypropyl) ether, 2,4,2',4'-tetrahydroxybenzophenone, and 2,4-dichlorophenoxyacetic acid. (Paper III) 4. We revealed the binding conformations of perfluorooctanesulfonic acid, perfluorooctanoic acid, and BP2 in the thyroxine binding sites (TBSs) of TTR by co-crystallizing TTR with the three compounds. A VS protocol was developed based on the TTR complex structures that predicted 192 industrial chemicals as potential binders to TTR. Seven novel TTR binders were confirmed by in vitro experiments including clonixin, 2,6-dinitro-p-cresol (DNPC), triclopyr, fluroxypyr, bisphenol S, picloram, and mesotrione. We further co-crystallized TTR with PBS, clonixin, DNPC, and triclopyr, and their complex structures showed that the compounds bind in the TBSs as proposed by the VS protocol. In summary, 13 indoor dust contaminants and industrial chemicals were identified as THDCs using a combination of in silico and in vitro approaches. To the best of our knowledge, none of these compounds has previously been reported to bind to TTR or THR. The identifications of these THDCs improve our understanding on the structure-activity relationships of THDCs. The crystal structures of TTR-THDC complexes and the information on THDC-Target intermolecular interactions provide a better understanding on the mechanism-of-actions behind thyroid disruption. The dataset compiled and in silico methods developed serve as a basis for identification of more diverse THDCs in the future and a tool for guiding de novo design of safer replacements.

付記する学位プログラム名: 充実した健康長寿社会を築く総合医療開発リーダー育成プログラム / 京都大学 / 0048 / 新制・課程博士 / 博士(薬科学) / 甲第22400号 / 薬科博第122号 / 新制||薬科||13(附属図書館) / 京都大学大学院薬学研究科医薬創成情報科学専攻 / (主査)教授 掛谷 秀昭, 教授 二木 史朗, 教授 土居 雅夫 / 学位規則第4条第1項該当 / Doctor of Pharmaceutical Sciences / Kyoto University / DFAM

Le développement des anticorps thérapeutiques s'est rapidement accéléré dans les 10 dernières années et concerne un nombre croissant de pathologies. La connaissance de l'épitope, à savoir la région de la cible à laquelle l'anticorps se fixe, est essentielle pour la compréhension des effets fonctionnels de ce dernier. Nous avons développé une méthode in silico, MAbTope, qui permet une prédiction précise de cet épitope, quand bien même aucune structure 3D de l'anticorps d’intérêt n'est résolue. Cette méthode se base sur une méthode d'amarrage protéine-protéine développée auparavant dans l’équipe BIOS. Le jeu d'apprentissage a été fortement enrichi en complexes anticorps-cibles, de nouvelles fonctions de score spécifiques ont été mises au point, et le plus important, l'objectif de l'apprentissage-machine a été modifié pour optimiser non plus la conformation de !'assemblage, mais la prédiction de l'épitope. Nous montrons que la méthode qui en résulte permet une prédiction précise et robuste de l'épitope, que la structure 3D de l'anticorps soit connue ou non. Nous montrons également comment les prédictions peuvent être facilement exploitées pour la validation expérimentale. Enfin, nous montrons comment la méthode peut être utilisée pour étudier à haut-débit le recouvrement d'épitopes par des anticorps ayant la même cible. / The development of therapeutic antibodies has been rapidly increasing in the last 10 years, with application to an increasing number of pathologies. The knowledge of the epitope, the region of the antigen to which the antibody binds, is crucial for understanding its functional effects. We have developed an in silico method, MAbTope, which allows the accurate prediction of the epitope, regardless of the availability of the 3D structure of the antibody of interest. This method is based on a protein-protein docking method previously developed in the BIOS group. The learning dataset was enlarged in antibody-antigen complexes, new specific scoring functions have been designed, and very importantly, the objective of machine-learning was switched from the conformational perspective towards the epitope determination perspective. We show that the resulting method allows robust and accurate prediction, whether or not the 3D structure of the antibody is available. We also show how the predictions can be easily exploited for experimental validation. Finally, we show how this method can be used for high-throughput epitope binning.