Global ETD Search

1	Application of theoretical methods to the study of small molecules in solution Lowis, D. R. January 1994 (has links) No description available. 536.7 Free energy perturbation theory
2	Advances in Ligand Binding Predictions using Molecular Dynamics Simulations Keränen, Henrik January 2014 (has links) Biochemical processes all involve associations and dissociations of chemical entities. Understanding these is of substantial importance for many modern pharmaceutical applications. In this thesis, longstanding problems with regard to ligand binding are treated with computational methods, applied to proteins of key pharmaceutical importance. Homology modeling, docking, molecular dynamics simulations and free-energy calculations are used here for quantitative characterization of ligand binding to proteins. By combining computational tools, valuable contributions have been made for pharmaceutically relevant areas: a neglected tropical disease, an ion channel anti-drug-target, and GPCR drug-targets. We report three compounds inhibiting cruzain, the main cysteine protease of the protozoa causing Chagas’ disease. The compounds were found through an extensive virtual screening study and validated with experimental enzymatic assays. The compounds inhibit the enzyme in the μM-range and are therefore valuable in further lead optimization studies. A high-resolution crystal structure of the BRICHOS domain is reported, together with molecular dynamics simulations and hydrogen-deuterium exchange mass spectrometry studies. This work revealed a plausible mechanism for how the chaperone activity of the domain may operate. Rationalization of structure-activity relationships for a set of analogous blockers of the hERG potassium channel is given. A homology model of the ion channel was used for docking compounds and molecular dynamics simulations together with the linear interaction energy method employed for calculating the binding free-energies. The three-dimensional coordinates of two GPCRs, 5HT1B and 5HT2B, were derived from homology modeling and evaluated in the GPCR Dock 2013 assessment. Our models were in good correlation with the experimental structures and all of them placed among the top quarter of all models assessed. Finally, a computational method, based on molecular dynamics free-energy calculations, for performing alanine scanning was validated with the A2A adenosine receptor bound to either agonist or antagonist. The calculated binding free-energies were found to be in good agreement with experimental data and the method was subsequently extended to non-alanine mutations. With extensive experimental mutation data, this scheme is a valuable tool for quantitative understanding of ligand binding and can ultimately be used for structure-based drug design. free-energy perturbation molecular dynamics ligand binding free-energy perturbation linear interaction energy binding free-energy homology modeling virtual screening alanine scanning amino acid mutagenesis hERG GPCR adenosine receptor serotonin receptor BRICHOS cruzain
3	From Structure to Function with Binding Free Energy Calculations for Codon Reading, Riboswitches and Lectins Sund, Johan January 2013 (has links) Molecular association is part of many important processes in living cells. Computational methods for calculating binding free energies allows for a quantitative examination of biomolecular structures and hypotheses drawn from biochemical experiments. Here, binding free energy calculations for tRNAs and release factors binding to mRNA codons on the ribosome, sugars binding to lectins and purine analogs binding to the purine riboswitch are presented. The relative affinities between cognate and non-cognate tRNAs for different states involved in codon reading on the ribosome were determined. The calculations show that tRNA discrimination varies between different conformations of the 30S subunit, where the existence of both low and high selectivity states provides an efficient common mechanism for initial selection and proofreading. The simulations reveal a desolvation mechanism for the 30S conformational switch with which the accuracy of peptide bond formation can be amplified. When an mRNA stop codon (UAA, UAG or UGA) is located in the ribosomal A-site release factors bind to the ribosome and the synthesized protein is released. RF1 is specific for UAA and UAG whereas RF2 is specific for UAA and UGA. The free energy calculations and an analysis of the performed simulations show the mechanisms for how RF1 and RF2 are able to read the stop codons with different specificities. Also mitochondrial release factors were investigated. Vertebrate mitochondria have four stop codons, UAA, UAG, AGA and AGG and two release factors mtRF1 and mtRF1a. The calculations show how the specificities of both mtRF1 and mtRF1a agree with RF1 and that none of them are likely to read the non-standard stop codons AGA and AGG. The linear interaction energy method has also been examined for the RSL and PA-IIL lectins and for the purine riboswitch. The standard parameterization of the method works well for RSL, but fails for PA-IIL and the purine riboswitch due to compositions of the active sites in these systems. The development of new parameterizations to overcome these problems leads to a better understanding of both the method and the binding mechanisms in these systems. binding free energy ribosome codon reading release factor mitochondrial translation purine riboswitch lectin molecular dynamics free energy perturbation
4	UNDERSTANDING CARBOHYDRATE RECOGNITION MECHANISMS IN NON-CATALYTIC PROTEINS THROUGH MOLECULAR SIMULATIONS Kognole, Abhishek A. 01 January 2018 (has links) Non-catalytic protein-carbohydrate interactions are an essential element of various biological events. This dissertation presents the work on understanding carbohydrate recognition mechanisms and their physical significance in two groups of non-catalytic proteins, also called lectins, which play key roles in major applications such as cellulosic biofuel production and drug delivery pathways. A computational approach using molecular modeling, molecular dynamic simulations and free energy calculations was used to study molecular-level protein-carbohydrate and protein-protein interactions. Various microorganisms like bacteria and fungi secret multi-modular enzymes to deconstruct cellulosic biomass into fermentable sugars. The carbohydrate binding modules (CBM) are non-catalytic domains of such enzymes that assist the catalytic domains to recognize the target substrate and keep it in proximity. Understanding the protein-carbohydrate recognition mechanisms by which CBMs selectively bind substrate is critical to development of enhanced biomass conversion technology. We focus on CBMs that target both oligomeric and non-crystalline cellulose while exhibiting various similarities and differences in binding specificity and structural properties; such CBMs are classified as Type B CBMs. We show that all six cellulose-specific Type B CBMs studied in this dissertation can recognize the cello-oligomeric ligands in bi-directional fashion, meaning there was no preference towards reducing or non-reducing end of ligand for the cleft/groove like binding sites. Out of the two sandwich and twisted forms of binding site architectures, twisted platform turned out to facilitate tighter binding also exhibiting longer binding sites. The exterior loops of such binding sites were specifically identified by modeling the CBMs with non-crystalline cellulose showing that high- and low-affinity binding site may arise based on orientation of CBM while interacting with non-crystalline substrate. These findings provide various insights that can be used for further understanding of tandem CBMs and for various CBM based biotechnological applications. The later part of this dissertation reports the identification of a physiological ligand for a mammalian glycoprotein YKL-40 that has been only known as a biomarker in various inflammatory diseases and cancers. It has been shown to bind to oligomers of chitin, but there is no known function of YKL-40, as chitin production in the human body has never been reported. Possible alternative ligands include proteoglycans, polysaccharides, and fibers such as collagen, all of which make up the mesh comprising the extracellular matrix. It is likely that YKL-40 is interacting with these alternative polysaccharides or proteins within the body, extending its function to cell biological roles such as mediating cellular receptors and cell adhesion and migration. We considered the feasibility of polysaccharides, including cello-oligosaccharides, hyaluronan, heparan sulfate, heparin, and chondroitin sulfate, and collagen-like peptides as physiological ligands for YKL-40. Our simulation results suggest that chitohexaose and hyaluronan preferentially bind to YKL-40 over collagen, and hyaluronan is likely the preferred physiological ligand, as the negatively charged hyaluronan shows enhanced affinity for YKL-40 over neutral chitohexaose. Collagen binds in two locations at the YKL-40 surface, potentially related to a role in fibrillar formation. Finally, heparin non- specifically binds at the YKL-40 surface, as predicted from structural studies. Overall, YKL-40 likely binds many natural ligands in vivo, but its concurrence with physical maladies may be related to the associated increases in hyaluronan. protein-carbohydrate interaction carbohydrate recognition glycosaminoglycan collagen molecular dynamics free energy perturbation Biochemical and Biomolecular Engineering Biochemistry Biophysics Structural Biology
5	Non-Steroidal Anti-Inflammatory Drugs in Cyclooxygenases 1 and 2 : Binding modes and mechanisms from computational methods and free energy calculations Shamsudin Khan, Yasmin January 2017 (has links) Non-steroidal anti-inflammatory drugs (NSAIDs) are one of the most commonly used classes of drugs. They target the cyclooxygenases (COX) 1 and 2 to reduce the physiological responses of pain, fever, and inflammation. Due to their role in inducing angiogenesis, COX proteins have also been identified as targets in cancer therapies. In this thesis, I describe computational protocols of molecular docking, molecular dynamics simulations and free energy calculations. These methods were used in this thesis to determine structure-activity relationships of a diverse set of NSAIDs in binding to their target proteins COX-1 and 2. Binding affinities were calculated and used to predict the binding modes. Based on combinations of molecular dynamics simulations and free energy calculations, binding mechanisms of sub-classes of NSAIDs were also proposed. Two stable conformations of COX were probed to understand how they affect inhibitor affinities. Finally, a brief discussion on selectivity towards either COX isoform is discussed. These results will be useful in future de novo design and testing of third-generation NSAIDs. molecular dynamics simulations binding free energy molecular docking cyclooxygenase non-steroidal anti-inflammatory drugs free energy perturbation potentials of mean force Pharmaceutical Biotechnology Läkemedelsbioteknik
6	Computational Modelling of Ligand Complexes with G-Protein Coupled Receptors, Ion Channels and Enzymes Boukharta, Lars January 2014 (has links) Accurate predictions of binding free energies from computer simulations are an invaluable resource for understanding biochemical processes and drug action. The primary aim of the work described in the thesis was to predict and understand ligand binding to several proteins of major pharmaceutical importance using computational methods. We report a computational strategy to quantitatively predict the effects of alanine scanning and ligand modifications based on molecular dynamics free energy simulations. A smooth stepwise scheme for free energy perturbation calculations is derived and applied to a series of thirteen alanine mutations of the human neuropeptide Y1 G-protein coupled receptor and a series of eight analogous antagonists. The robustness and accuracy of the method enables univocal interpretation of existing mutagenesis and binding data. We show how these calculations can be used to validate structural models and demonstrate their ability to discriminate against suboptimal ones. Site-directed mutagenesis, homology modelling and docking were further used to characterize agonist binding to the human neuropeptide Y2 receptor, which is important in feeding behavior and an obesity drug target. In a separate project, homology modelling was also used for rationalization of mutagenesis data for an integron integrase involved in antibiotic resistance. Blockade of the hERG potassium channel by various drug-like compounds, potentially causing serious cardiac side effects, is a major problem in drug development. We have used a homology model of hERG to conduct molecular docking experiments with a series of channel blockers, followed by molecular dynamics simulations of the complexes and evaluation of binding free energies with the linear interaction energy method. The calculations are in good agreement with experimental binding affinities and allow for a rationalization of three-dimensional structure-activity relationships with implications for design of new compounds. Docking, scoring, molecular dynamics, and the linear interaction energy method were also used to predict binding modes and affinities for a large set of inhibitors to HIV-1 reverse transcriptase. Good agreement with experiment was found and the work provides a validation of the methodology as a powerful tool in structure-based drug design. It is also easily scalable for higher throughput of compounds. computer simulations molecular dynamics ligand binding free energy perturbation linear interaction energy binding free energy homology modelling structure prediction alanine scanning site-directed mutagenesis hERG GPCR neuropeptide Y HIV-1 reverse transcriptase integron integrase
7	Computational Studies of Protein Synthesis on the Ribosome and Ligand Binding to Riboswitches Lind, Christoffer January 2017 (has links) The ribosome is a macromolecular machine that produces proteins in all kingdoms of life. The proteins, in turn, control the biochemical processes within the cell. It is thus of extreme importance that the machine that makes the proteins works with high precision. By using three dimensional structures of the ribosome and homology modelling, we have applied molecular dynamics simulations and free-energy calculations to study the codon specificity of protein synthesis in initiation and termination on an atomistic level. In addition, we have examined the binding of small molecules to riboswitches, which can change the expression of an mRNA. The relative affinities on the ribosome between the eukaryotic initiator tRNA to the AUG start codon and six near-cognate codons were determined. The free-energy calculations show that the initiator tRNA has a strong preference for the start codon, but requires assistance from initiation factors 1 and 1A to uphold discrimination against near-cognate codons. When instead a stop codon (UAA, UGA or UAG) is positioned in the ribosomal A-site, a release factor binds and terminates protein synthesis by hydrolyzing the nascent peptide chain. However, vertebrate mitochondria have been thought to have four stop codons, namely AGA and AGG in addition to the standard UAA and UAG codons. Furthermore, two release factors have been identified, mtRF1 and mtRF1a. Free-energy calculations were used to determine if any of these two factors could bind to the two non-standard stop codons, and thereby terminate protein synthesis. Our calculations showed that the mtRF’s have similar stop codon specificity as bacterial RF1 and that it is highly unlikely that the mtRF’s are responsible for terminating at the AGA and AGG stop codons. The eukaryotic release factor 1, eRF1, on the other hand, can read all three stop codons singlehandedly. We show that eRF1 exerts a high discrimination against near-cognate codons, while having little preference for the different cognate stop codons. We also found an energetic mechanism for avoiding misreading of the UGG codon and could identify a conserved cluster of hydrophobic amino acids which prevents excessive solvent molecules to enter the codon binding site. The linear interaction energy method was used to examine binding of small molecules to the purine riboswitch and the FEP method was employed to explicitly calculate the LIE b-parameters. We show that the purine riboswitches have a remarkably high degree of electrostatic preorganization for their cognate ligands which is fundamental for discriminating against different purine analogs. Binding free energy Ribosome Codon reading Translation initiation Translation termination Mitochondrial translation Release factor Purine riboswitch Molecular Dynamics Free Energy Perturbation Biochemistry and Molecular Biology Biokemi och molekylärbiologi Bioinformatics (Computational Biology) Bioinformatik (beräkningsbiologi)
8	Compréhension de l'énantiosélectivité de la lipase B de Candida antarctica : étude par modélisation moléculaire et expérimentation / Comprehensive study of Candida antarctica lipase B enantioselectivity : using experimental and molecular modeling approaches Chaput, Ludovic 28 September 2012 (has links) La lipase B de Candida antarctica (CALB) est un enzyme présentant des propriétés énantiosélectives très intéressantes pour l’obtention de molécules énantio pures par dédoublement cinétique de mélanges racémiques,molécules utilisées comme synthons dans l’industrie pharmaceutique. En effet, le principe actif de nombreux médicaments est efficace sous une forme énantio pure, l’autre forme chirale pouvant se révéler délétère pour l’organisme.Les travaux de la thèse s’intéressent à mieux comprendre l’origine de l’énantiosélectivité de la lipase B de Candida antarctica, en particulier pour la résolution d’alcools secondaires par des réactions de transestérification.Nous utilisons pour la première fois la méthode de la perturbation de l’énergie libre pour estimer la différence d’énergie libre entre les intermédiaires tétraédriques obtenus avec les formes R et S d’alcools énantiomères pour une série d’alcools secondaires, dans le but de prédire in silico l’énantiosélectivité de la CALB. Les paramètres cinétiques apparents d’une réaction avec deux alcools substrats énantiopurs sont expérimentalement déterminés et permettent de définir la contribution respective du Km et du kcat de chaque énantiomère pour la définition de l’énantiosélectivité. L’étude expérimentale de l’effet d’empreinte par des molécules co-substrats est réalisée,ainsi qu’une étude par modélisation moléculaire de l’effet d’empreinte par le premier ester substrat de la réaction qui pourrait modifier la conformation du site actif de la CALB. La troisième partie porte sur l’étude de la CALB et de trois variants (T42V, S47A et T42V/S47A) chez lesquels les acides aminés dans la poche stéréospécifiques ont mutés. T42V et S47A permettent d’obtenir une augmentation de l’énantiosélectivité. L’étude propose une étude détaillée de la conformation du site actif à partir de simulations de trajectoires de dynamique moléculaire / The lipase B from Candida antarctica is an enzyme displaying enantioselective properties which are interesting to obtain enantio pure compounds by kinetic resolution of racemic mixtures, which are used as pharmaceutical intermediates and fine chemicals. Indeed, for most of the drugs, only one of the two chiral formsis efficient as bioactive compound, whereas the other chiral form may display deleterious effects. Present work concerns the understanding of the origin of Candida antarctica lipase B enantioselectivity, and more especially in case of the resolution of secondary alcohols by transesterification. We used, for the first time, the free energy perturbation method to evaluate the free energy difference between tetrahedral intermediates with R and S alcohol enantiomers for a series of secondary alcohols in order to predict in silico enantiomeric ratio of CALB-catalyzed reactions. The apparent kinetic parameters were experimentally determined for two enantio pure substrates and allow to evalute the relative contribution of both Km and kcat for R and S enantiomers in the enantiomeric ratio of CALB-catalyzed reactions. Experimental study of imprinting effect hypothesis by co-substrate molecules was done. Molecular modeling studies of imprinting effect hypothesis were performed, in which the first substrate ester of the reaction could mould the active site. At least, the third part of this thesis concerns the study of wild-type CALB and three different variants (T42V and S47A which allow to increase enantioselectivity and T42/S47A) of CALB by molecular modeling. A detailed study of the conformation of the stereo specificity pocket in the active site is presented, based on molecular dynamics simulations. Lipase B de Candida antarctica Énantiosélectivité Biocatalyse Modélisation moléculaire Intermédiaire tétraédrique Perturbation de l’énergie libre Effet d’empreinte Mutant T42V S47A T42V/S47A Lipase B from Candida antarctica Enantioselectivity Biocatalysis Molecular modeling Tetrahedral intermediate Free energy perturbation Imprinting effect Mutant T42V S47A T42V/S47A
9	Electrostaticanalisys the Ras active site Khan, Abdul Kareem 05 March 2009 (has links) La preorganització electrostàtica del centre actiu s'ha postulat com el mecanisme genèric de l'acció dels enzims. Així, alguns residus "estratègics" es disposarien per catalitzar reaccions interaccionant en una forma més forta amb l'estat de transició, baixant d'aquesta manera el valor de l'energia dactivació g cat. S'ha proposat que aquesta preorientació electrostática s'hauria de poder mostrar analitzant l'estabilitat electrostàtica de residus individuals en el centre actiu.Ras es una proteïna essencial de senyalització i actúa com un interruptor cel.lular. Les característiques estructurals de Ras en el seu estat actiu (ON) són diferents de les que té a l'estat inactiu (OFF). En aquesta tesi es duu a terme una anàlisi exhaustiva de l'estabilitat dels residus del centre actiu deRas en l'estat actiu i inactiu. / The electrostatic preorganization of the active site has been put forward as the general framework of action of enzymes. Thus, enzymes would position "strategic" residues in such a way to be prepared to catalyze reactions byinteracting in a stronger way with the transition state, in this way decreasing the activation energy g cat for the catalytic process. It has been proposed thatsuch electrostatic preorientation should be shown by analyzing the electrostatic stability of individual residues in the active site.Ras protein is an essential signaling molecule and functions as a switch in thecell. The structural features of the Ras protein in its active state (ON state) are different than those in its inactive state (OFF state). In this thesis, an exhaustive analysis of the stability of residues in the active and inactive Ras active site is performed. dinàmica centre actiu relacions d'estructura activitat preorganització reorganització estat de transició estat actiu estabilitat electrostàtica interacción proteïna-proteïna test de ranks de Wilcoxon P-loop G1 motif HVR metastability RMSD substrate assisted catalysis sequence alignment beta sheet helix Z-test wilcoxon rank test protein-protein interactions electrostatic stability active state dynamics signal transduction electrostatic stabilization ras effectors guanine exchange factor GTPase-activating proteins quantum mechanics/molecular PDLD/S-LRA X-ray crystallography solvation energy free energy perturbation statistical analysis GTP hydrolysis guanosine diphosphate guanosine triphosphate computer simulation thermodynamics protein conformation electrostatics interruptor II interruptor I llaç P motiu G1 HVR (regions altament variables) metaestabilita RMSD catàlisi assistida pel substrat aliniament de seqüència fulla beta hèlix test Z transducció del senyal estabilització electrostàtica efectors de ras factor de bescamvi de guanina proteïnes activadores de l'activitat mecànica quàntica/mecànica PDLD/S-LRA cristalografia de raigs X energia de solvatació perturbació de l'energia lliure anàlisi estadística hidròlisi de GTP difosfat de guanosina trifosfat de guanosina simulació computacional termodinàmica conformació proteïca electrostàtica switch-I switch-II 537 576

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