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CHARACTERIZING THE BINDING INTERACTION BETWEEN DICYANOGOLD (I) AND HUMAN SERUM ALBUMINMoore, Alison Blythe January 2002 (has links)
No description available.
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Understanding Amyloid Inhibition: Toward a Residue-Resolution Map of the Interactions between the Alzheimer's Aβ-Peptide and Human Serum AlbuminAlgamal, Moustafa 11 1900 (has links)
Amyloidogenesis refers to a process of protein misfolding and aggregation that leads to the formation of highly stable amyloid fibers. Amyloidogenesis may lead to loss of physiological protein function and/or formation of toxic intermediates, which are linked to mutliple human diseases. Amyloidogenesis is inhibited by plasma proteins, which function as extracellular chaperones by binding to stressed and misfolded proteins, including amyloidogenic peptides, and preventing their aggregation. This thesis focuses on the ability of human serum albumin (HSA), the main protein in human plasma, to inhibit amyloidogenesis, with emphasis on the molecular nature of the interactions between HSA and the amyloid β peptide (Aβ) associated with Alzhemier’s disease. HSA is as a key amyloidogenic regulator, a novel function for this protein that goes beyond the traditional HSA roles as plasma osmotic pressure regulator and as binder and carrier of endogenous and exogenous low molecular weight ligands. As a first step towards understanding the detailed molecular nature of these interactions, this thesis will focus on defining the key binding determinants in the interaction between HSA and Aβ peptides. Primarily, we will try to answer two main questions. First, which HSA residues are critical for the recognition of Aβ peptides and the prevention of Aβ aggregation? Second, which Aβ residues are mostly affected by HSA binding? Starting form our knowledge about the stoichiometry and affinity of the Aβ interactions at the level of HSA domains, Chapter 2 addresses the first question through successful applications of a reductionist approach, based on a combination of mutational comparative analyses and fatty acid (FA) competition. This strategy allowed us to identify a short HSA derived peptide that specifically recognizes Aβ and prevents its aggregation. In Chapter 3, we examine the effect of HSA on the pseudo-equilibrium state between Aβ monomers and protofibrils. Using Dark state Exchange Saturation Transfer (DEST), Saturation Transfer Difference (STD) and 15N T2 relaxation experiments, we show that Aβ peptides interact with HSA via a dual mechanism. First, selected residues in Aβ (1-40) monomers bind specifically but weakly to HSA (Kd = 0.1 - 1 mM). Second, HSA competes with Aβ monomers for the binding to the protofibrils, as indicated by an HSA-dependent decrease in the direct vs. tethered probabilities for contacts between Aβ monomer residues and the protofibril surface. The effect of HSA mimics that of dilution for the majority of the Aβ (1-40) residues involved in the cross-beta strands of amyloid fibrils. Finally, Chapter 4 will outline future investigations to address currently open questions about HSA dynamics, HSA-Aβ and HSA-FA interactions, for which we acquired preliminary data. / Thesis / Master of Science (MSc)
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Universal Aqueous-Based Antifouling Coatings for Multi-Material DevicesGoh, Sharon January 2017 (has links)
Biofouling is an ongoing problem in the development and usage of biomaterials for biomedical implants, microfluidic devices, and water-based sensors. Antifouling coatings involving surface modification of biomaterials is widely utilized to reduce unwanted protein adsorption and cell adhesion. Surface modification strategies, however, are reliant on the working material’s chemical properties. Thus, published procedures are often not applicable to a wide range of material classes. This constitutes a serious limitation in using surface modification on assembled multi-material devices, i.e on whole device modification. The objective of this research is to develop an antifouling coating with non-aggressive reaction conditions that can universally modify polymers and other material classes. Two strategies using polydopamine (PDA) as an anchor for polyethylene glycol (PEG) surface attachment were investigated: (1) PDA-PEG backfilled with bovine serum albumin (BSA), and (2) PDA-PEG with light activated perfluorophenyl azide (PFPA) conjugated to the PEG. Three materials varying in surface wettability were studied to evaluate the coatings for multi-material applications: porous polycarbonate membrane (PC), polydimethyl siloxane (PDMS), and soda lime glass cover slips.
Atomic force microscopy (AFM) and ellipsometry studies revealed substantial structural differences of PDA. Differences in PDA surface roughness affected PEG grafting in solution (the first method), with higher PEG coverage achieved on PC with intermediate surface roughness to PDMS and glass. Radiolabeled Fg adsorption and E. coli adhesion experiments showed reduced fouling on all PDA-PEG modified materials when backfilled with BSA. The ability for BSA to penetrate the PEG layer indicated that low PEG grafting densities were achieved using this grafting-to approach. The use of a photoactive labeling agent, PFPA, to tether PEG was proposed to improve PEG grafting on PDA. The PFPA-PEG modification protocol was optimized by quantifying Fg adsorption. Two treatments of PFPA-PEG were required to fully block PDA active sites. Fg adsorption was not significantly improved on PFPA-PEG modified PC and glass when backfilled with BSA, indicating sufficient PEG coverage of PDA. High Fg adsorption on PFPA-PEG surfaces indicate that high density PEG brushes were still not achieved with this method. PDMS surfaces were damaged with this procedure due to increased surface handling in the protocol. This is the first, to our knowledge, successful demonstration of PFPA modification on PDA surfaces. Photopatterning of polymer-based materials can be achieved, providing opportunities for utilising new materials in cell patterned platforms. Due to low PEG coverage on PDA surfaces from solution and using PFPA, ultra-low protein adsorption cannot be achieved using these aqueous-based methods. Antifouling modifications using PDA and PEG should be applied for short-term cell studies. / Thesis / Master of Applied Science (MASc)
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Microbubble fermentation of recombinant Pichia pastoris for human serum albumin productionZhang, Wei 24 July 2003 (has links)
The high cell density fermentation of recombinant Pichia pastoris for human serum albumin (HSA) production is a high oxygen demand process. The oxygen demand is usually met by increased agitation rate and use of oxygen-enriched air. Microbubble fermentation however can supply adequate oxygen to the microorganisms at relatively low agitation rates because of improved mass transfer of the microbubbles used for the sparging. Conventionally sparged fermentations were conducted for the production of HSA using P. pastoris at agitation rates of 350, 500, and 750 rpm, and were compared to MBD sparged fermentation at 150, 350, and 500 rpm agitation rates. The MBD improved the volumetric oxygen transfer coefficient (kLa) and subsequently increased the cell mass and protein production compared to conventional fermentation.
Cell production in MBD fermentation at 350 rpm was 4.6 times higher than that in conventional fermentation at 350 rpm, but similar to that in the conventional 750 rpm. Maximum cell mass productivity in the conventional 350 rpm was only 0.37 g / (L·h), while the maximum value in MBD 350 rpm was 2.0 g / (L·h), which was similar to 2.2 g / (L·h) in the conventional 750 rpm. Biomass yield on glycerol Ys (g cell/ g glycerol) was 0.334 g / g in the conventional 350 rpm, 0.431 g / g in MBD 350 rpm and 0.438 g / g in the conventional 750 rpm. Protein production in MBD 350 rpm was 7.3 times higher than that in the conventional 350 rpm, but similar to the conventional 750 rpm. Maximum protein productivity in the conventional 350 rpm was 0.37 mg / (L·h), 2.8 mg / (L·h) in MBD 350 rpm, and 3.3 mg / (L·h) in the conventional 750 rpm. Protein yield on methanol Yp (mg protein / g methanol) was 1.57 mg /g in the conventional 350 rpm, 5.02 in MBD 350 rpm, and 5.21 in the conventional 750 rpm.
The volumetric oxygen transfer coefficient kLa was 1011.9 h-1 in MBD 350 rpm, which was 6.1 times higher than that in the conventional 350 rpm (164.9 h-1) but was similar to the conventional 750 rpm (1098 h-1). Therefore, MBD fermentation results at low agitation of 350 rpm were similar to those in the conventional fermentation at high agitation of 750 rpm. There was considerable improvement in oxygen transfer to the microorganism using MBD sparging relative to the conventional sparging.
Conventional fermentations were conducted both in a Biostat Q fermenter (small) at 500 rpm, 750 rpm, and 1000 rpm, and in a Bioflo III fermenter (large) at 350 rpm, 500 rpm, and 750 rpm. At the same agitation rate of 500 rpm, cell production in the large reactor was 3.8 times higher than that in the small one, and no detectable protein was produced in the small reactor at 500 rpm. At the same agitation rate of 750 rpm, both cell production and protein production in the large reactor were 4.6 times higher than the small reactor. Thus, the Bioflo III fermenter showed higher oxygen transfer efficiency than the Biostat Q fermenter, because of the more efficient aeration design of the Bioflo III fermenter. / Master of Science
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Studies on the use of bovine serum albumin as aerosol performance enhancer in dry powder inhalation formulations prepared by spray drying. / 小牛血清白蛋白(BSA)對以噴霧乾燥(spray dry)制作的粉霧吸入劑(DPI)粉霧性能(aerosol performance)提升的研究 / Xiao niu xue qing bai dan bai (BSA) dui yi pen wu qan zao (spray dry) zhi zuo de fen wu xi ru ji (DPI) fen wu xing neng (aerosol performance) ti sheng de yan jiuJanuary 2010 (has links)
Chan, Pui. / "November, 2009." / Thesis (M.Phil.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (leaves 108-114). / Abstracts in English and Chinese. / Table of Contents --- p.i / Acknowledgement --- p.vi / Abstract --- p.vii / Abstract (Chinese) --- p.ix / Chapter Chapter One --- Introduction / Chapter 1.1. --- Pulmonary Route for Drug Delivery --- p.2 / Chapter 1.2. --- Factors Affecting the Performance of Inhaled Formulations --- p.3 / Chapter 1.2.1. --- Particle Aerodynamic Diameter --- p.4 / Chapter 1.2.2. --- Dispersibility of Particles --- p.5 / Chapter 1.2.3. --- Clearance Mechanism in Lung and Dissolution of Particles --- p.6 / Chapter 1.3. --- Production of Dry Powder Inhalation by Spray Drying --- p.7 / Chapter 1.4. --- Approaches to Enhance Aerosol Performance of Spray Dried Particles --- p.8 / Chapter 1.4.1 --- Porous/Hollow Particles --- p.9 / Chapter 1.4.2 --- Non-Porous Corrugated Particles --- p.10 / Chapter 1.4.3 --- Blends and Ternary Systems --- p.10 / Chapter 1.4.4 --- Surface Energy and Crystallinity Modification --- p.11 / Chapter 1.4.5 --- Other Approaches to Enhancing Aerosol Performance --- p.12 / Chapter 1.5 --- Objectives and Rationale of the Present Study --- p.13 / Chapter 1.6 --- Scope of Present Study and Particle Characterization Techniques Employed --- p.14 / Chapter 1.6.1 --- Microscopy and Particle Density Measurements --- p.14 / Chapter 1.6.2 --- Particle Size Analysis and Particle Dispersibility --- p.15 / Chapter 1.6.3 --- Thermal Analysis and Particle Crystallinity --- p.15 / Chapter 1.6.4 --- Particle Surface Characterization --- p.16 / Chapter 1.6.5 --- Inverse Gas Chromatography --- p.18 / Chapter 1.6.6 --- Fractal Analysis --- p.19 / Chapter 1.6.6.1 --- Background and Origin of Fractal Analysis --- p.19 / Chapter 1.6.6.2 --- Use of Fractal Analysis in Pharmaceutical Research --- p.20 / Chapter 1.6.6.3 --- Methods for fractal analysis --- p.21 / Chapter 1.6.7 --- Atomic Force Microscopy --- p.23 / Chapter 1.6.7.1 --- Background of Atomic Force Microscopy --- p.23 / Chapter 1.6.7.2 --- Characterization of Surface Topography by Atomic Force Microscopy --- p.23 / Chapter 1.6.7.3 --- Measurement of Interaction Forces by Colloid Probe 226}0Ø Microscopy --- p.25 / Chapter 1.6.7.4 --- Use of Atomic Force Microscopy in Pharmaceutical Research --- p.27 / Chapter Chapter Two --- Materials and Methods / Chapter 2.1. --- Materials --- p.30 / Chapter 2.2. --- Equipment --- p.31 / Chapter 2.3. --- Methods --- p.33 / Chapter 2.3.1. --- Powder Preparation --- p.33 / Chapter 2.3.1.1 --- Preparation of Salbutamol Sulphate Samples --- p.33 / Chapter 2.3.1.2 --- Preparation of Disodium Cromoglycate Samples --- p.33 / Chapter 2.3.1.3 --- Preparation of ß-Galactosidase (BG) Samples --- p.34 / Chapter 2.3.2. --- Determination of Aerosol Performance --- p.35 / Chapter 2.3.3. --- Determination of Protein Activity for BG Samples --- p.36 / Chapter 2.3.3.1. --- Enzyme Assay Procedure --- p.37 / Chapter 2.3.3.2. --- Calculation of Enzyme Activity --- p.38 / Chapter 2.3.3.3. --- Determination of Enzyme Activity Retained in Spray-dried Samples --- p.38 / Chapter 2.3.4. --- Physicochemical Characterization of Particles --- p.39 / Chapter 2.3.4.1. --- Scanning Electron Microscopy --- p.39 / Chapter 2.3.4.2. --- Particle Density Determination --- p.39 / Chapter 2.3.4.3. --- Particle Size Analysis --- p.40 / Chapter 2.3.4.4. --- Thermal analysis --- p.41 / Chapter 2.3.4.5. --- Powder X-ray Diffraction --- p.42 / Chapter 2.3.4.6. --- Surface Area Determination --- p.42 / Chapter 2.3.4.7. --- Surface Composition Characterization --- p.43 / Chapter 2.3.4.8. --- Surface Tension Measurement --- p.44 / Chapter 2.3.4.9. --- Inverse Gas Chromatography --- p.45 / Chapter 2.3.4.9.1. --- Calculation of Standard Free Energy of Adsorption --- p.46 / Chapter 2.3.4.9.2. --- Calculation of Dispersive Component of Surface Free Energy --- p.47 / Chapter 2.3.4.9.3. --- Determination of Specific Interactions and Associated Acid-Base Properties --- p.48 / Chapter 2.3.4.10. --- Fractal Analysis --- p.49 / Chapter 2.3.4.11. --- Atomic Force Microscopy --- p.49 / Chapter Chapter Three --- Results / Chapter 3.1. --- In vitro Aerosol Performance --- p.52 / Chapter 3.2. --- Enzyme Activity Retained in BG Samples --- p.55 / Chapter 3.3. --- Scanning Electron Microscopy (SEM) --- p.56 / Chapter 3.3.1. --- SEM of Salbutamol Sulphate Formulations --- p.56 / Chapter 3.3.2. --- SEM of DSCG Formulations --- p.59 / Chapter 3.3.3. --- SEM of BG Formulations --- p.61 / Chapter 3.4. --- Density Measurements --- p.65 / Chapter 3.4.1. --- Densities of Salbutamol Sulphate Formulations --- p.65 / Chapter 3.4.2. --- Densities of DSCG Formulations --- p.66 / Chapter 3.4.3. --- Densities of BG Formulations --- p.67 / Chapter 3.5. --- Particle Size Analysis by Laser Diffraction --- p.68 / Chapter 3.5.1. --- Volume Mean Diameter Measurements --- p.68 / Chapter 3.5.2. --- Particle Size Distributions and Dispersion Patterns of Formulations --- p.70 / Chapter 3.6. --- Thermal Analysis --- p.75 / Chapter 3.7. --- Powder X-ray Diffraction --- p.80 / Chapter 3.8. --- Surface Area Measurements --- p.84 / Chapter 3.9. --- Surface Composition Characterization --- p.85 / Chapter 3.9.1. --- Surface Composition of Salbutamol Sulphate Formulations --- p.85 / Chapter 3.9.2. --- Surface Composition of DSCG Formulations --- p.88 / Chapter 3.9.3. --- Surface Composition of BG/BSA Formulations --- p.89 / Chapter 3.10. --- Surface Tension Measurements --- p.91 / Chapter 3.11. --- Inverse Gas Chromatography --- p.92 / Chapter 3.12. --- Fractal Analysis --- p.93 / Chapter 3.13. --- Atomic Force Microscopy --- p.94 / Chapter Chapter Four --- Discussion / Chapter 4.1. --- Influence of BSA on Aerosol Performance and Protein Integrity --- p.98 / Chapter 4.2. --- Influence of BSA on Physicochemical Properties of Particles --- p.98 / Chapter 4.2.1. --- Influence of BSA on surface corrugation --- p.98 / Chapter 4.2.2. --- Influence of BSA on particle size and dispersion behavior --- p.99 / Chapter 4.2.3. --- Influence of BSA on crystallinity and thermal properties of particles --- p.100 / Chapter 4.2.4. --- Influence of BSA on surface energetics of particles --- p.100 / Chapter 4.3. --- Relationship between Surface Corrugation and Aerosol Performance --- p.101 / Chapter 4.4. --- Mechanism of Surface Modification for BSA on Spray-dried Particles --- p.103 / Chapter Chapter Five --- Conclusions and Future Work / Chapter 5.1. --- Conclusions --- p.106 / Chapter 5.1.1. --- General Aerosolization-Enhancing Effect of BSA --- p.106 / Chapter 5.1.2. --- Surface Modifying Effect of BSA --- p.106 / Chapter 5.1.3. --- Relationship between Surface Corrugation and Aerosol Performance --- p.106 / Chapter 5.2. --- Future Work --- p.107 / References --- p.108
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Protein interaction with polyelectrolytes and ligands: A structural and thermodynamic investigationYu, Shun 29 August 2017 (has links)
Der erste Teil dieser Arbeit untersucht die Ladungswechselwirkung zwischen Proteinen und Polyelektrolyten. Dabei wird die Bindung von Polyakrylsäure (PAA) als kurzes Modell-Polyelektrolyt an Humanalbumin (HSA) in einer umfassenden experimentellen und theoretischen Studie untersucht und sehr gute Übereinstimmung der Resultate konnte festgestellt werden. Die Computersimulationen in dieser Arbeit wurden von Xiao Xu im Rahmen seiner Promotion durchgeführt. Thermodynamische Daten wurden mit Hilfe von Isothermer Titrationskalorimetrie (ITC) gesammelt und strukturelle Untersuchungen wurden mit Hilfe von Neutronenkleinwinkelstreuung (SANS) durchgeführt. Die Analyse von Bindungsaffinitäten zeigte eine eins zu eins Bindung von PAA mit HSA, die entropisch getrieben ist und strukturellen Untersuchungen weisen eine stabile Proteilstruktur unabhängig von der Adsorption durch PAA auf.
Im zweiten Teil der Arbeit wird die Wechselwirkung zweier uremischer Toxinen, nämlich Phenylessigsäure (PhAA) und Indoxylsulfat (IDS), mit HSA untersucht. Eine wichtige Schlussfolgerung aus der Analyse der ITC Daten ist, dass begünstigende, hydrophobe Wechselwirkungen die treibende Kraft für die Adsorption von Toxinen an HSA sind, und dass hierbei die Enthalpie-Entropie-Kompensation (EEC) zu tragen kommt. Weiterhin zeigen SANS Untersuchungen, dass die Proteinstruktur trotz Adsorption stabil bleibt und konnte außerdem über Interpartikulare Wechselwirkung von HSA-Toxin Komplexen aufklären.
Im Allgemeinen ist HSA strukturell unverändert durch die Adsorption von Liganden. Diese Feststellung erlaubt die Interpretation von ITC Daten, da damit gemessene Wärmeprozesse ausschließlich von Bindungsprozessen herrühren. Die vorliegende Arbeit konnte zeigen, dass eine ausführliche thermodynamische Analyse durch Kombination von theoretischer mit experimenteller Arbeit, eine umfassende Einsicht in die Mechanismen von Bindungsprozessen ermöglicht. / The first part of the thesis explores the charge-charge interaction between proteins and polyelectrolytes. Polyacrylic acid (PAA) is used as a short model polyelectrolyte to interact with human serum albumin (HSA) the most abundand protein in blood, in a comprehensive experimental and theoretical study. The results thereby coincide very well. Computer simulation studies were performed by Xiao Xu within the framework of his PhD thesis. Thermodynamic data were collected by means of isothermal titration calorimetry (ITC) and structural analysis performed using small-angle neutron scattering (SANS). The analysis of binding free energies revealed one to one binding that is mainly driven by entropy. Structural investigations give proof of the stability of the protein beside adsorption.
In the second part, the interaction of two uremic toxins, namely phenylacetic acid (PhAA) and indoxyl sulfate (IDS), with HSA is studied. Systematic ITC experiments reveal two binding sites for both of the two toxins and show small dependence of binding affinities on ionic strength in contrast to PAA adsorption to HSA. This leads to the key conclusion that the favorable hydrophobic interaction is the driving contribution for adsorption and enthalpy-entropy compensation (EEC) effect comes into play. SANS studies of high concentrated HSA-toxin solutions proofed the stability of the protein structure and shed light on the interparticle interaction of HSA-toxin complexes.
In general, HSA is structurally robust regardless of ligand uptake. This finding allows the interpretation of ITC data by confirming that the measured heat signals are purely associated to the binding process. The present thesis has demonstrated that a full thermodynamic analysis in combination with theoretical modelling can provide a comprehensive understanding of binding in terms of identifying driving forces and their contributions to protein ligand interaction.
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Computational and micro-analytical techniques to study the in vitro and in silico models of novel therapeutic drugsGumede, Njabulo Joyfull January 2016 (has links)
Submitted in fulfillment of the requirements for the Doctor of Philosophy degree in Chemistry, Durban University of Technology, Durban, South Africa, 2016. / In drug discovery and development projects, metabolism of new chemical entities (NCEs) is a major contributing factor for the withdrawal of drug candidates, a major concern for other chemical industries where chemical-biological interactions are involved. NCEs interact with a target macro-molecule to stimulate a pharmacological or toxic response, known as pharmacodynamics (PD) effect or through the Adsorption, Distribution, Metabolism, and Excretion (ADME) process, triggered when a bio-macromolecule interacts with a therapeutic drug. Therefore, the drug discovery process is important because 75% of diseases known to human kind are not all cured by therapeutics currently available in the market. This is attributed to the lack of knowledge of the function of targets and their therapeutic use in order to design therapeutics that would trigger their pharmacological responses.
Accordingly, the focus of this work is to develop cost saving strategies for medicinal chemists involved with drug discovery projects. Therefore, studying the synergy between in silico and in vitro approaches maybe useful in the discovery of novel therapeutic compounds and their biological activities. In this work, in silico methods such as structure-based and ligand-based approaches were used in the design of the pharmacophore model, database screening and flexible docking methods. Specifically, this work is presented by the following case studies:
The first involved molecular docking studies to predict the binding modes of catechin enantiomer to human serum albumin (HSA) interaction; the second involved the use of docking methods to predict the binding affinities and enantioselectivity of the interaction of warfarin enantiomers to HSA. the third case study involved a combined computational strategy in order to generate information on a diverse set of steroidal and non-steroidal CYP17A1 inhibitors obtained from literature with known experimental IC50 values. Finally, the fourth case study involved the prediction of the site of metabolisms (SOMs) of probe substrates to Cytochrome P450 metabolic enzymes CYP 3A4, 2D6, and 2C9 making use of P450 module from Schrödinger suite for ADME/Tox prediction.
The results of case study I were promising as they were able to provide clues to the factors that drive the synergy between experimental kinetic parameters and computational thermodynamics parameters to explain the interaction between drug enantiomers and thetarget protein. These parameters were correlated/converted and used to estimate the pseudo enantioselectivity of catechin enantiomer to HSA. This approach of combining docking methodology with docking post-processing methods such as MM-GBSA proved to be vital in estimating the correct pseudo binding affinities of a protein-ligand complexes. The enantioselectivity for enantiomers of catechin to HSA were 1,60 and 1,25 for site I and site II respectively.
The results of case study II validates and verifies the preparation of ligands and accounting for tautomers at physiological pH, as well as conformational changes prior to and during docking with a flexible protein. The log KS = 5.43 and log KR = 5.34 for warfarin enantiomer-HSA interaction and the enantioselectivity (ES = KS/KR) of 1.23 were close to the experimental results and hence referred to as experimental-like affinity constants which validated and verified their applicability to predict protein-ligand binding affinities.
In case study III, a 3D-QSAR pharmacophore model was developed by using 98 known CYP17A1 inhibitors from the literature with known experimental IC50 values. The starting compounds were diverse which included steroidal and non-steroidal inhibitors. The resulting pharmacophore models were trained with 69 molecules and 19 test set ligands. The best pharmacophore models were selected based on the regression coefficient for a best fit model with R2 (ranging from 0.85-0.99) & Q2 (ranging from 0.80-0.99) for both the training and test sets respectively, using Partial Least Squares (PLS) regression. On the other hand, the best pharmacophore model selected was further used for a database screening of novel inhibitors and the prediction of their CYP17A1 inhibition. The hits obtained from the database searches were further subjected to a virtual screening workflow docked to CYP17A1 enzyme in order to predict the binding mode and their binding affinities. The resulting poses from the virtual screening workflow were subjected to Induced Fit Docking workflow to account for protein flexibility during docking. The resulting docking poses were examined and ranked ordered according to the docking scores (a measure of affinity).
Finally, the resulting hits designed from an updated model from case study III were further synthesized in an external organic chemistry laboratory and the synthetic protocols as well as spectroscopic data for structure elucidation forms part of the provisional patent specification. A provisional patent specification has been filed (RSA Pat. Appln. 2015/ 07849). The case studies performed in this thesis have enabled the discovery of non-steroidal CYP17A1 inhibitors. / D
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MRI kontrastní látky pro angiografické aplikace / MRI contrast agents for angiographyUrbanovský, Peter January 2015 (has links)
Modern diagnostic method magnetic resonance imaging (MRI) usually uses contrast agents T1-type, which are based on Gd3+ complexes. Due to severe toxicity of free Gd3+ , it is desired to have thermodynamically stable and kinetically inert complexes with fast elimination from the body. This work summarizes information about a novel contrast agent based on ligand DO3AP (1,4,7,10-tetraazacyclododecane-1-methyl(alkyl)phosphinic-4,7,10- triacetic acid) with pendant hydrophobic dibenzylamino group which is able to interact hydrophobically with the macromolecule of serum albumin. The stability of supracomplex is dependent on pH value, i.e. on the protonation of the pendant amino group of the complex (pKA = 5.6) and this interaction was confirmed from 1 H-NMRD profile and fluorescent analysis. The compound was tested for its angiographic properties in vivo on rat model. Furthermore, other complexes of the ligand with trivalent lanthanides (Nd3+ , Eu3+ , Tb3+ , Dy3+ , Er3+ ) were characterized by various methods (XRD, luminescence, UV-VIS, 1 H-, 17 O- and 31 P-NMR). The cleavage of the benzyl groups affords ligand whose Ln3+ complexes possess pH dependent PARACEST effect. These complexes were characterized by XRD, luminescence and 1 H- and 31 P-NMR. Moreover, the novel ligands with modified length of pendant...
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Investigação química de complexos de coordenação dos antibióticos enrofloxacina e norfloxacina combinados ao íon Ru(III) e suas interações com biomoléculas alvo / Chemical Investigation of coordination compounds with enrofloxacin and norfloxacin antibiotics combined to Ru (III) ion and their interations with target biomolecule.Reis, Felipe Costa Claro 28 July 2014 (has links)
Este trabalho tem como objetivo sintetizar e caracterizar um novo complexo mononuclear de rutênio (III) e enrofloxacina (enro, fármaco antibacteriano da família das fluoroquinolonas), [Ru(enro)3].nH2O. Foram testadas várias rotas sintéticas e apenas a partir de uma delas obteve-se o composto desejado. O produto foi caracterizado pelas técnicas espectroscópicas de absorção na região do UV-visível e do infra-vermelho. Através desta última técnica foi possível determinar o modo pelo qual a enrofloxacina se coordena ao íon rutênio: a coordenação ocorre de modo bidentado através do oxigênio da piridona e do oxigênio do grupamento carboxilato. Outro objetivo deste trabalho foi investigar a interação do complexo mononuclear de rutênio (III) e norfloxacina, [Ru(nor)3].nH2O, com a albumina de soro humano (HSA), através da técnica de luminescência. Mais especificamente pelo estudo da supressão da luminescência dos resíduos de triptofano, aplicando-se o modelo de tratamento da supressão bimolecular de Stern-Volmer. O estudo de supressão de fluorescência mostrou, por meio de espectros de emissão da HSA, que com o aumento da concentração do complexo [Ru(nor)3].nH2O na solução de HSA, ocorre uma redução gradual da luminescência da HSA, devido a alterações da conformação da proteína, que sugerem alteração do microambiente próximos aos resíduos de triptofano. A partir do tratamento dos dados pode-se determinar tanto K_sv quanto a constante cinética do processo de supressão, que mostraram uma dependência com a temperatura sugerindo como mecanismo predominante de supressão o mecanismo dinâmico. Porém essa conclusão foi revista a partir da determinação dos tempos de vida do estado excitado da HSA, e pode-se concluir que o mecanismo predominante à temperatura ambiente é o mecanismo estático, porém com o aumento da temperatura ocorre a predominância do mecanismo do tipo dinâmico. Através da determinação dos parâmetros termodinâmicos, concluiu-se que as interações entre a HSA e o complexo são espontâneas, e forças de van der Waals e ligações de hidrogênio estão envolvidas na ligação entre a HSA e o supressor. / This work aims to synthesize and characterize a new mononuclear ruthenium (III) complex and enrofloxacin (enro, antibacterial drug of the fluoroquinolone family), [Ru(enro)3].nH2O. Several synthetic routes were tested, but only from one of them it was obtained the desired compound. The product was characterized by spectroscopic techniques of absorption in UV-visible and infra-red regions. Through this last technique, it was possible to determine the coordination mode of enrofloxacin to the ruthenium ion: the coordination occurs in a bidentate way through the pyridone oxygen and the oxygen of the carboxylate group. Another aim of this study was to investigate the interaction of mononuclear ruthenium (III) complex and the norfloxacin, [Ru(nor)3].nH2O, with the human serum albumin (HSA), through the technique of luminescence. More specifically, by the study of the quenching of luminescence of tryptophan residues, by applying the Stern-Volmers model of treatment of bimolecular suppression. The fluorescence quenching study showed, through the emission spectra of HSA, that increasing the complex concentration in HSA solution, there is a gradual reduction of the luminescence of HSA, due to the conformational changes of the protein that suggests the change of microenvironment near tryptophan residues. From the data processing it is possible to determine both K_sv and the kinetic constant of the suppression process, which showed temperature dependence, suggesting as the predominant mechanism of quenching the dynamic mechanism. However, this conclusion has been revised from the determination of the lifetimes of the excited state of HSA, and it can be concluded that the predominant mechanism at room temperature is the static mechanism, but with the temperatures increase, it occurs the predominance of the dynamic type mechanism. By determining the thermodynamic parameters, it was concluded that the interactions between HSA and the complex are spontaneous, and Van der Waals forces and hydrogen bonds are involved in the binding between HSA and suppressor.
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Caracterização do aço inoxidável austenítico UNS S31254 em meio de NaCI 0,11 mol L-1 visando seu emprego em implantes ortopédicos / Electrochemical characterization of UNS S31254 austenitic stainless steel in 0.11 mol L-1 NaCl medium in order to propose its application in orthopaedic implantsAfonso, Monica Luisa Chaves de Andrade 27 September 2006 (has links)
Foi feita a caracterização eletroquímica do aço inoxidável austenítico UNS S31254 em meio de NaCl 0,11 mol L-1 na ausência e presença de soro albumina bovina (BSA) visando seu emprego em implantes ortopédicos. Foram empregadas como técnicas: medidas de potencial de circuito aberto, curvas de polarização, cronoamperometria, EIE, XPS, MEV, EDS e EEO. O comportamento eletroquímico do aço 254 foi comparado com o de outros aços empregados em implantes ortopédicos (ISO 5832-9, ASTM F138, e AISI 316L) na ausência e presença de BSA. O aço 254 se mostrou semelhante ao ISO 5832-9: encontra-se passivado desde o potencial de corrosão até o de transpassivação; a presença de inclusões de óxidos de cálcio e alumínio no aço 254 foi considerada a responsável por um potencial de transpassivação 100 mV menos positivo do que o observado com o aço ISO 5832-9. Foi detectada. além de óxido de Cr(III), a presença de Mo na forma Mo(VI) no filme passivo do aço 254. A ação da BSA, ora passivante ora catalisadora, depende de sua concentração, da natureza do substrato metálico, e do potencial na interfase metal-solução. A BSA modifica o mecanismo de oxidação do aço 254 e inibe seletivamente a dissolução dos seus elementos constituintes, em particular, níquel e cromo. / The electrochemical characterization of UNS S31254 has been made in 0.11 mol L-1 NaCl medium in the absence and presence of bovine serum albumin (BSA) in order to propose its application in orthopaedic implants. The techniques employed were: open circuit potential measurements, polarization curves, chronoamperometry, EIS, XPS, SEM, EDS and EEO. The electrochemical behavior of 254 SS was compared to that observed for ISO 5832-9, ASTM F138 and AISI 316L stainless steels, used in orthopedic implants, in the absence and presence of BSA. 254 SS is similar to ISO 5832-9 SS: it is passivated on the potential range between the corrosion and the transpassivation potentials; the presence of calcium and aluminum oxides can be responsible for the shift of about 100 mV to less positive potentials on the transpassivation potential when compared to ISO 5832-9 SS. The presence of Mo(VI) was detected beside Cr(III) as passivating film for 254 SS. BSA action depends on its concentration, the nature of the metallic substract and on the potential in the metal-solution interphase. BSA changes the oxidation mechanism of 254 SS and promotes the selective dissolution of the elements particularly nickel and chromium.
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