This work was aimed at developing novel tools that utilize HINT, an empirical forcefield capable of quantitating both hydrophobic and hydrophilic (hydropathic) interactions, for implementation in theoretical biology and drug discovery/design. The role of hydrophobicity in determination of macromolecular structure and formation of complexes in biological molecules is undeniable and has been the subject of research across several decades. Hydrophobicity is introduced, with a review of its history and contemporary theories. This is followed by a description of various methods that quantify this all-pervading phenomenon and their use in protein folding and contemporary drug design projects – including a detailed overview of the HINT forcefield. The specific aim of this dissertation is to introduce our attempts at developing new methods for use in the study of antibacterial drug resistance and antiviral drug discovery. Multidrug efflux is commonly regarded as a fast growing problem in the field of medicine. Several species of microbes are known to have developed resistance against almost all classes of antibiotics by various modes-of-action, which include multidrug transporters (a.k.a. efflux pumps). These proteins are present in both gram-positive and gram-negative bacteria and extrude molecules of various classes. They protect the efflux pump-expressing bacterium from harmful effects of exogenous agents by simply evacuating the latter. Perhaps the best characterized mechanism amongst these is that of the AcrA-AcrB-TolC efflux pump. Data is available in literature and perhaps also in proprietary databases available with pharmaceutical companies, characterizing this pump in terms of the minimum inhibitory concentration ratios (MIC ratios) for various antibiotics. We procured a curated dataset of 32 β-lactam and 12 antibiotics of other classes from this literature. Initial attempts at studying the MIC ratios of β-lactam antibiotics as a function of their three dimensional topology via 3D-quantitative structure activity relationship (3D-QSAR) technology yielded seemingly good models. However, this methodology is essentially designed to address single receptor-ligand interactions. Molecules being transported by the efflux pump must undoubtedly be involved in multiple interactions with the same. Notably, such methods require a pharmacophoric overlap of ligands prior to the generation of models, thereby limiting their applicability to a set of structurally-related compounds. Thus, we designed a novel method that takes various interactions between antibiotic agents and the AcrA-AcrB-TolC pump into account in conjunction with certain properties of the drugs. This method yielded mathematical models that are capable of predicting high/low efflux with significant efficiency (>93% correct). The development of this method, along with the results from its validation, is presented herein. A parallel aim being pursued by us is to discover inhibitors for hemagglutinin-neuraminidase (HN) of human parainfluenza virus type 3 (HPIV3) by in silico screening. The basis for targeting HN is explored, along with commentary on the methodology adopted during this effort. This project yielded a moderate success rate of 34%, perhaps due to problems in the computational methodology utilized. We highlight one particular problem – that of emulating target flexibility – and explore new avenues for overcoming this obstacle in the long run. As a starting point towards enhancing the tools available to us for virtual screening in general (and for discovering antiviral compounds in specific), we explored the compatibility between sidechain rotamer libraries and the HINT scoring function. A new algorithm was designed to optimize amino acid residue sidechains, if provided with the backbone coordinates, by generating sidechain positions using the Dunbrack and Cohen backbone-dependent rotamer library and scoring them with the HINT scoring function. This rotamer library was previously used by its developers previously to design a very successful sidechain optimization algorithm called SCWRL. Output structures from our algorithm were compared with those from SCWRL and showed extraordinary similarities as well as significant differences, which are discussed herein. This successful implementation of HINT in our sidechain optimization algorithm establishes the compatibility between this forcefield and sidechain rotamer libraries. Future aims in this project include enhancement of our current algorithm and the design of a new algorithm to explore partial induced-fit in targets aimed at improving current docking methodology. This work shows significant progress towards the implementation of our hydropathic force field in theoretical modeling of biological systems in order to enhance our ability to understand atomistic details of inter- and intramolecular interactions which must form the basis for a wide variety of biological phenomena. Such efforts are key to not only to understanding the said phenomena, but also towards a solid basis for efficient drug design in the future.

Computationally driven drug design/discovery efforts generally rely on accurate assessment of the forces that guide the molecular recognition process. HINT (Hydropathic INTeraction) is a natural force field, derived from experimentally determined partition coefficients that quantifies all non-bonded interactions in the biological environment, including hydrogen bonding, electrostatic and hydrophobic interactions, and the energy of desolvation. The overall goal of this work is to apply the HINT-based atomic level description of molecular systems to biologically important proteins, to better understand their biochemistry – a key step in exploiting them for therapeutic purposes. This dissertation discusses the results of three diverse projects: i) structural modeling of human sphingosine kinase 2 (SphK2, a novel anticancer target) and binding mode determination of an isoform selective thiazolidine-2,4-dione (TZD) analog; ii) structural modeling of human cytomegalorvirus (HCMV) alkaline nuclease (AN) UL98 (a novel antiviral target) and subsequent virtual screening of its active site; and iii) explicit treatment of interfacial waters during protein-protein docking process using HINT-based computational tools. SphK2 is a key regulator of the sphingosine-rheostat, and its upregulation /overexpression has been associated with cancer development. We report structural modeling studies of a novel TZD-analog that selectively inhibits SphK2, in a HINT analysis that identifies the key structural features of ligand and protein binding site responsible for isoform selectivity. The second aim was to build a three-dimensional structure of a novel HCMV target – AN UL98, to identify its catalytically important residues. HINT analysis of the interaction of 5’ DNA end at its active site is reported. A parallel aim to perform in silico screening with a site-based pharmacophore model, identified several novel hits with potentially desirable chemical features for interaction with UL98 AN. The majority of current protein-protein docking algorithms fail to account for water molecules involved in bridging interactions between partners, mediating and stabilizing their association. HINT is capable of reproducing the physical and chemical properties of such waters, while accounting for their energetic stabilizing contributions. We have designed a solvated protein-protein docking protocol that explicitly models the Relevant bridging waters, and demonstrate that more accurate results are obtained when water is not ignored.

Children’s right to participation in situations that matter to them is stated in law and convention texts and is emphasized by the children themselves in research studies, too. When actively involved, their perspective is visualized. Children’s use of interactive technology has increased considerably during the last decade. The use of applications and web sites are becoming a regular occurrence in paediatric healthcare. The overall aim was to develop and test, together with children, an interactive communication tool meant to facilitate young children’s participation in healthcare situations. To understand children’s varied perceptions of their involvement in healthcare situations, interviews, drawings and vignettes were used in a phenomenographic approach (I). A participatory design iteratively evaluated evolving prototypes of an application (II). Video observations and hermeneutics captured the meanings of the participation cues that the children demonstrated when they used the application in healthcare situations (III). A quantitative approach was used to identify patterns in the children’s cue use (IV). In total, 114 children in two clinical settings and in a preschool were involved. The result showed that the children perceived themselves, their parents and the professionals as actors in a healthcare situation, although all were perceived to act differently (I). The children contributed important information on age-appropriateness, usability and likeability in the iterative evaluating phases that eventually ended up in the application (II). When using the application in healthcare situations, the cues they demonstrated were understood as representing a curious, thoughtful or affirmative meaning (III). Curious cues were demonstrated to the highest extent. The three-year-olds and the children with the least experience of healthcare situations demonstrated the highest numbers of cues (IV). Conclusion: when using the application, the children demonstrated a situated participation which was influenced by their perspective of the situation and their inter-inter-action with the application as well as the health professional. The children’s situated participation provided the professionals’ with additional ways of guiding the children based on their perspectives. / I lag- och konventionstexter, liksom i forskning, som använder barnens egna uttryck betonas deras rättighet att vara delaktiga i situationer av betydelse för dem. Genom att delta kan de göra sitt perspektiv synligt. Under det senaste årtiondet har barns användning av interaktiv teknik ökat kraftigt. Applikationer och web-sidor används nuförtiden även flitigt inom barnsjukvården. Avhandlingens övergripande syfte var att utveckla och pröva, tillsammans med barn, ett interaktivt kommunikationsverktyg avsett att möjliggöra yngre barns delaktighet i vårdsituationer. Intervjuer, teckningar och vignetter användes för att, fenomenografiskt, förstå barnens uppfattningar av att vara i en vårdsituation (I). En iterativ deltagarbaserad design användes för att utveckla en prototyp av en applikation. En hermeneutisk tolkning av video-observationer fångade meningen i barnens sätt att visa sin delaktighet (hintar) vid användningen av applikationen i vårdsituationer (III). En deduktiv, kvantitativ ansats användes för att identifiera mönster i barnens sätt att visa sin delaktighet när de använde applikationen (IV). Resultatet visade att barnen uppfattade sig själva, föräldrarna och vårdpersonalen som aktörer i situationen, även om alla uppfattades agera på olika sätt (I). Barnen bidrog i den iterativa processen med viktig information gällande aspekter som åldersanpassning, användbarhet och hur väl den tilltalar dem, vilket slutligen ledde fram till den färdiga applikationen (II). Barnens sätt att visa sin delaktighet när de använde applikationen förstods ha en nyfiken, tankfull och självbekräftande mening (III). Nyfikenheten visades mest vid användningen av applikationen. Treåringarna samt barnen med minst vårderfarenhet använde applikationen i störst utsträckning (IV). Sammanfattning: när applikationen användes i vårdsituationen visade barnen en situerad delaktighet vilken byggde på deras perspektiv på den aktuella situationen samt på deras inter-inter-aktion med applikationen och vårdpersonalen. Genom detta erbjöds vårdpersonalen ytterligare ett sätt att guida barnet utifrån barnets eget perspektiv.

Ce travail porte sur les relatives dans les langues de l’aire Caucase – Anatolie de l’Est – Iran de l’Ouest dans une perspective aréale. Cette aire est connue pour concentrer une importante diversité génétique et typologique, avec une cinquantaine de langues appartenant à six familles distinctes : caucasique du Nord-Est, caucasique du Nord-Ouest, caucasique du Sud, turcique, indo-européenne et sémitique. Un grand nombre de ces langues sont en situation de contact et peuvent potentiellement s’influencer les unes les autres. Les relatives constituent une caractéristique intéressante à étudier dans une telle perspective car les constructions ayant fonction de relative peuvent varier selon nombreux paramètres d’une langue à l’autre ou au sein d’une même langue (position de la relative, finitude du verbe, etc.) ; chaque valeur possible pour un paramètre donné étant un trait pouvant potentiellement se diffuser. La première partie de cette thèse consiste en une description détaillée des stratégies de relativisation des langues de l’aire. Diverses stratégies sont identifiées, dont certaines couramment mentionnées dans la littérature, mais d’autres également plus atypiques du point de vue typologique. La deuxième partie consiste en une analyse des données décrites dans la première partie : après avoir identifié de manière précise les paramètres de variation et leurs différentes valeurs possibles parmi les stratégies disponibles de l’aire (environ 40 paramètres, 80 sous-paramètres et plus de 200 valeurs possibles), plusieurs zones géographiques sont mises en évidence où une même stratégie ou une même valeur pour un paramètre donné sont partagées par des langues non nécessairement apparentées génétiquement. Pour plusieurs de ces zones, il semble s’agir d’un phénomène de convergence, étant donné que les langues concernées divergent de leurs familles génétiques respectives et/ou le trait ou la stratégie partagé(e) est atypique. Pour finir, plusieurs facteurs pouvant intervenir dans la diffusion ou non d’une stratégie ou d’un trait linguistique sont suggérés. / This work deals with relative clauses in an areal perspective, focusing on languages of Caucasus – Eastern Anatolia – Western Iran area. This area is known to display an important linguistic diversity (genetic as well as typological) with about fifty languages belonging to six distinct families: North-East Caucasian, North-West Caucasian, South-Caucasian, Turkic, Indo-European and Semitic. Most of these languages are in contact and may influence each other. Besides, relative clauses constitute an interesting field to investigate in an areal perspective, given that the constructions used as relative clauses may vary according to numerous parameters, crosslinguistically or within the same language (e.g., position of the relative clause, finiteness of the verbal form, etc.); each value for a given parameter is a feature that may possibly diffuse across languages.The first part of this work is a fine-grained description of the relativization strategies available in the area. Various strategies are identified. The area displays all strategies commonly mentioned in the literature, as well as some typologically less common strategies.The second part of the work is an analysis of the data. After identifying all variation parameters and their different values in the strategies available in the area (i.e., about 40 main variation parameters, 80 sub-parameters and more than 200 potential values), several geographical areas are highlighted where the same strategy or the same value for a given parameter is shared by languages that are not necessarily genetically related. For some of these areas, it seems that a convergence phenomena is involved given that languages may diverge with this respect from their respective genealogical families and/or the shared feature or strategy is typologically uncommon. Finally, some factors that could be involved in the occurrence or not of a language-contact induced change are suggested. / Bu tezde bölgesel bir açıdan Kafkas – Doğu Anadolu – Batı İran bölgesinde konuşulan dillerde sıfat işlevli yan tümceler inceleniyor. Hem genetik hem de tipolojik olarak dilsel zenginliği ile tanınan bu bölgede günümüzde altı farklı aileden (Kuzeydoğu Kafkas dilleri, Kuzeybatı Kafkas dilleri, Güney Kafkas dilleri, Türk dilleri, Hint-Avrupa dilleri, Sami dilleri) elliyi aşkın dil konuşulmaktadır. Bu dillerin çoğu birbirleri ile temas halindeler ve birbilerini etkileyebiliyorlar. Bu bakımdan, sıfat işlevli yan tümce olarak kullanılan yapıların diller arasında, hatta aynı dil içinde dahi birçok parametreye göre çeşitlilik gösterdiğini (örneğin yan tümcenin nitelediği ada göre konumu, yan tümcedeki fiil şekli (tam donanımlı ya da sıfat-fiil oluşu), vb.), ve bu parametrelerin alabileceği değerlerin bir dilden diğerine yayılabilen bir nitelikte olduğunu göz önüne alırsak, sıfat işlevli yan tümcelerin bölgesel ve karşılaştırmalı bir açıdan ele alınabilecek zengin bir çalışma konusu olduğu belirgin hale gelmektedir. Tezin ilk kısmında, adı geçen bölgede konuşulan dillerdeki sıfat işlevli yan tümce yapıları ayrıntılı bir şekilde belgeleniyor. Bu belgeleme literatürde sıkça bahsi geçen alışılmış yapıların yanı sıra çok bilinmeyen, atipik yapıları da gün yüzüne çıkarıyor.Tezin ikinci kısmında ise birinci kısımda belgelenen veriler çözümleniyor. Sıfat işlevli yan tümcelere ait tüm parametreleri ve bu parametrelerin alabileceği farklı değerleri (toplamda yaklaşık 40 ana parametre, 80 alt-parametre ve 200’den fazla olası değer) saptadıktan sonra, aynı yapıyı ya da belli bir parametreye ait aynı değeri kullanan fakat genetik açıdan birbiri ile ilişkisi olmayan dillerin oluşturduğu birtakım coğrafi alan tespit ediliyor. Bu alanların bazısında saptanan diller, kullandıkları sıfat işlevli yan tümce yapıları veya bu yapılardaki bazı parametrelerin aldığı değerler bakımından genetik olarak ilişkili oldukları dillerden farklılık gösterdiklerinden ve/veya kullandıkları yapıların tipolojik olarak alışılmış olmadıklarından dolayı, bu belirtilen alanlarda bir yöneşme (convergence) meydana gelme olasılığı yüksektir. Son olarak da bir yapının ya da belli bir parametre değerinin yayılıp yayılmamasına sebep olabilecek bir kaç etken öneriliyor.