A successful cancer treatment is a combination of early diagnosis and efficient use of anticancer drugs. There is a chance of approximately 70 - 90% of cancer patients surviving if the diagnosis is conducted early. That means if a diagnosis system is in place which can detect multiple types of cancer at an early stage, a potential cancer therapy is most likely to succeed. However, at present, the available biomedical sensors are unable to detect and differentiate between cancerous cells or tumours. They are also not able to provide continuous real-time monitoring of a patient. Additionally, oral anticancer drugs given during chemotherapy, at the moment, suffer from low bioavailability. Also, a variety of these drugs is not targeted in nature. That means the drug will potentially affect areas of the body which do not need it. The low bioavailability of the drug will not only increase the chemotherapy sessions but also makes the entire process more aggravating for the cancer patient. Therefore, there is an absolute need to have innovative and efficient anticancer drug delivery mechanisms. Finally, current biomedical sensors are primarily made up of silicon (Si) or hard substrates based materials. Even if the biomedical sensor is of a flexible material, the material is either a fragile film or flexible but not stretchable polymers such as polyimide (PI). By having a biomedical sensor which is moderately flexible or not flexible at all, a continuous on-body biomedical sensing is not possible in an efficient manner. That is because hard substrates based biomedical sensors would be difficult to be placed on a body at all times. Furthermore, the flexible biomedical sensors currently suffer from problems such as the electrode on top cracking and damaging after few uses rendering them unusable. Hence, a new fabrication process needs to be devised to solve the issues mentioned above. In this work, an attempt is made to utilise zinc oxide (ZnO) nanostructures for biomedical sensing, drug delivery and biomimetics. ZnO nanostructures are synthesised by using a low-cost wet chemistry process known as hydrothermal growth. Due to the inherent biocompatibility and unique electrical/ piezoelectric properties of ZnO, they acted as prime candidates for the applications outlined above. A high-throughput process is used to synthesise ZnO nanowires (NWs) on Si, polyimide-onsilicon (PI/Si) and directly on PI and polydimethylsiloxane (PDMS) substrates. The work utilises a variety of characterization tools. ZnO nanostructures' morphology is characterised by using a Scanning Electron Microscope (SEM), Transmission Electron Microscope (TEM) and Atomic Force Microscope (AFM). X-ray diffraction (XRD) was used to calculate the crystallite size and the crystalline orientation of the nanostructures. A novel fabrication process is developed to allow direct synthesis and direct patterning of metal electrodes on fully flexible, stretchable and bendable PDMS substrates by using standard photolithography. This novel fabrication process makes the PDMS substrates not expand when exposed to temperatures up to 110 °C. Also; the new fabrication process does not cause the PDMS to swell when exposed to various chemicals such as isopropyl alcohol (IPA) or acetone. The fabrication process has created a new paradigm shift in the field of patterning and producing devices directly on flexible and stretchable substrates. The PDMS substrate is further utilised as a sensitive bovine serum albumin (BSA) protein sensor which is capable of detecting up to femtomolar concentrations in just under 5 min of incubation time. Protein biosensing tests were carried out by measuring the change in resistance at 1V bias voltage. The PDMS based biosensor is tested as a protein sensor because proteins are important biomarkers in cancer diagnosis. Also, protein sensors are immensely useful in the detection of bacteria and viruses thereby allowing further expansion to the technology developed herewith. For the first time, ZnO NWs are used to deliver hydrophobic organic dye, Nile red, in a human body like environment. The Nile red simulates an anticancer drug as they share similar surface chemistry. There is an approximately 80% release of Nile red which shows that ZnO NWs can be used as an efficient anticancer drug delivery system with high bioavailability. For the drug delivery experiments, the dynamic dialysis based release of Nile red (Nr) from the ZnO nanowires is carried out by using UV-Visible (UV-Vis) spectroscopy. Fourier Transform Infrared (FTIR) was used to determine the coordination of Nr across the ZnO nanowires. Finally, a novel synthesis process is used to produce individual ZnO NWs on a single ZnO nanoplate (NP) which are named as ZNWNP nanostructures. ZNWNP nanostructures have high hydrophobicity without the need of any functionalization. The hydrophobicity of the hybrid ZnO nanowires on ZnO nanoplate nanostructures (ZNWNP) is characterised by using contact angle goniometry (CAG). Various contact angle theories have been used to calculate the surface free energy (SFE) of the ZNWNP nanostructures. The high hydrophobicity allows these nanostructures to be used for biomimetic applications such self-cleaning, bioinspired sensors and multimodal biosensing. Additionally, ZNWNP nanostructures can be used in biomedical sensors to create multimodal analysis. The multimodal analysis is immensely useful in cancer detection as at least three or more cancer biomarkers can be used to triangulate the diagnosis. The work presented in the thesis aims to utilise ZnO nanostructures for a variety of biomedical applications. The new fabrication process mentioned above has applications not only in biomedicine but also in the flexible electronics industry. The biomimetic nanostructures combined with the biomedical sensor gives rise to a robust multimodal analysis system which can change the course of the cancer diagnosis. That coupled with the usage of ZnO NWs as an effective anticancer drug delivery system gives an immense promise in advancing cancer therapy as a whole and making the entire treatment process less aggravating and less painful for cancer patients.

Thesis (M.Sc.Eng.) / This research addressed the design and fabrication of nano-patterned sapphire substrates (NPSS) as well as the growth by molecular-beam epitaxy (MBE) on such substrates of AlGaN and InGaN multiple quantum wells (MQWs). In recent years a number of LED manufacturers are developing nitride LED devices emitting in the visible part of the electromagnetic spectrum on micron-patterned sapphire substrate (MPSS). These devices are reported to have lower threading dislocation densities, resulting in improvement of the LED internal quantum efficiency (IQE). Furthermore, the LED devices fabricated on MPSS were also found to have improved light extraction efficiency (LEE), due to light scattering by the patterned substrate. My research focuses on the development of nano-patterned sapphire substrate aiming to improve the performance of LEDs grown by MBE and emitting at the deep ultraviolet region of the electromagnetic spectrum. In order to optimize the nano-patterning of the sapphire substrates for maximum light-extraction, the Finite-Difference Time-Domain (FDTD) simulation method was employed. The LEE enhancement was calculated as a function of the diameter, height and perion of the pattern. The calculations were performed only at a single wavelength, corresponding to the maximum of the emitted LED spectrum, which was taken to be 280 nm. These calculations have shown that the best sapphire substrate patterning strategy for this wavelength is the cone shape pattern in hexagonal array structure. Based on limited number of calculations I found that the optimum period, diameter and height of this cone shaped pattern are 400nm 375nm and 375nm respectively. Experimentally, nano patterned substrates were fabricated through natural and nano-imprint lithography. In natural lithography the first step for the definition of the nano-pattern consists of coating the sapphire substrate with photoresist (PMMA) followed by depositing a monolayer of polystyrene nanospheres, 400nm in diameter, using the Langmuir–Blodgett method. These spheres assemble on the substrate and form a closed packed hexagonal array pattern. Following this step the size of the spheres was slightly reduced using reactive-ion etching (RIE) in oxygen plasma. This was followed by the deposition a chromium film, lift-off to remove the polystyrene spheres and RIE to remove the PMMA from the footprints of the spheres. The substrate was then coated with a nickel or chromium films followed by another lift-off which defines the final mask for the formation of cone shaped features by RIE in a CHF3 plasma. An alternative method for pattern definition was the nanoimprint lithography; the stamp for this method (2 mm2 in size) was formed on Silicon substrate using e-beam lithography. NPSS with high quality pillar shape was also fabricated by this method, however, this method can produce only small size patterns. AlGaN films and GaN/InGaN MQWs were deposited on the NPSS by MBE, and characterized by Scanning electron microscopy and photoluminescence and cathodoluminescence measurements. The cathodoluminescence and photoluminescence spectra show that films grown on NPSS has much stronger luminescence than the films grown on flat sapphire substrate, consistent with enhanced light extraction efficiency.

Noble metal nanomaterials have numerous uses in plasmonic and surface enhanced Raman scattering (SERS) detection applications; however, upon the addition of analytes, nanomaterials often undergo uncontrolled aggregation which leads to inconsistent signal intensities. To overcome this limitation, the effect of gold nanosphere concentration, column purification, and surface chemistry functionalization using internally etched silica stabilization methods was investigated on SERS assays for small molecule detection. Nanostructure composition, size, shape, stability, surface chemistry, optical properties, and SERS-activity were monitored using localized surface plasmon resonance (LSPR or extinction) spectroscopy, transmission electron microscopy (TEM), and Raman spectroscopy. First, the behavior of citrate-stabilized gold nanospheres was monitored as a function of molecular surface coverage. Both extinction and SERS spectral intensities increased linearly below monolayer functionalization. Above this value, however, uncontrolled nanoparticle aggregation occurred and large but irreproducible SERS signal intensities were monitored. Next, gold nanoparticles were encapsulated with varying silica shell thicknesses and purified using traditional centrifugation steps and/or column chromatography. Relative to the traditionally purified (i.e. centrifuged) samples, the SERS responses from small molecules using the column purified nanoparticle samples followed a well-known SERS distance-dependence model. Thus, surface chemistry cannot form more than a 2 nm thick layer on gold nanospheres if SERS applications were targeted. To overcome these challenges, gold nanospheres encapsulated with a thick silica shell were made SERS-active by etching the internal silica layer near the metal surface. During the synthesis of these internally etched silica-coated gold nanospheres, the LSPR wavelength shift, a parameter related to the effective local refractive index near the gold core, was monitored instead of etching time, in order to produce nanostructures with more uniform internal silica etching from sample to sample. The SERS-activity of a target molecule using these nanostructures was measured as a function of LSPR wavelength shift. SERS signal intensity increased, which suggested that more analyte molecules were able to bind to the gold surface because of the larger pore size in the silica layer near the metal core. Further exploration of these findings should increase the integration of solution-phase nanoparticles in more predictable functions in future applications, resulting in more quantitative and reproducible molecular detection in complex sample matrices, including biological and environmental samples.

In this thesis, a number of commercially available paper substrates of various types are characterized and their characteristics related to the performance of inkjet-printed conductors using silver nanoparticle ink. The evaluated performance variables are electrical conductivity as well as the minimum achievable conductor width and the edge raggedness. It is shown that quick absorption of the ink carrier is beneficial for achieving well defined conductor geometry and high conductivity. Surface roughness with topography variations of sufficiently large amplitude and frequency is detrimental to print definition and conductivity. Porosity is another important factor, where the characteristic pore size is much more important than the total pore volume. A nearly ideal porous coating has large total pore volume but small characteristic pore size, preferably smaller than individual nanoparticles in the ink. Apparent surface energy is important for non-absorbing substrates but of limited importance for coatings with a high absorption rate.Additionally, a concept for improving the geometric definition of inkjet-printed conductors on nonporous films has been demonstrated. By coating the films with polymer–based coatings to provide a means of ink solvent removal, minimum conductor width were reduced a factor 2 or more.Intimately connected to the end performance of printed conductors is a well adapted sintering methodology. A comparative evaluation of a number of selective sintering methods has been performed on paper substrates with different heat tolerance. Pulsed high-power white light was found to be a good compromise between conductivity performance, reliability and production adaptability.The purpose of the work conducted in this thesis is to increase the knowledge base in how surface characteristics of papers and flexible films affect performance of printed nanoparticle structures. This would improve selection, adaption of, or manufacturing of such substrates to suit printed high conductivity patterns such as printed antennas for packaging. / I denna avhandling har ett antal kommersiellt tillgängliga papper av olika typ karaktäriserats och deras egenskaper relaterats till prestandan på inkjet-tryckta elektriska ledare tryckta med silvernanopartikelbläck. De undersökta prestandavariablerna är elektrisk ledningsförmåga samt ledarnas minimala linjebredd och kantjämnhet. Det visas att en snabb absorption av bläckets lösningsmedel är gynnsam för både väldefinierad ledningsgeometri och elektrisk ledningsförmåga. Ytråhet med topografiska variationer med tillräckligt stor amplitud och spatiell frekvens korrelerar negativt med tryckdefinition och ledningsförmåga. Porositet är ytterligare en viktig faktor, där karaktäristisk porstorlek är avsevärt viktigare än total porvolym. Nära ideala egenskaper hos en porös bestrykning synes vara en mycket hög total porvolym men med små individuella porer, med fördel mindre än de minsta metallpartiklarna i bläcket. Ytenergi är mycket betydelsefull för icke-absorberande substrat men tappar nästan all sin betydelse för bestrykningar med snabb absorption.Ett koncept för att förbättra den geometriska definitionen på inkjet-tryckta ledare på icke-porösa flexibla filmer har visats. Genom att bestryka filmerna med vissa polymerbaserade material och därmed införa en mekanism för separering av lösningsmedel och partiklar så reducerades ledarnas minimibredd med en faktor 2 eller mer.Intimt förknippad med den slutliga elektriska prestandan på tryckta ledare är också en väl anpassad sintringsmetodik. En jämförande utvärdering av ett flertal selektiva sintringmetoder har genomförts på papper med olika värmetålighet. Pulsat vitt ljus med hög effekt bedömdes som en bra kompromiss mellan elektriska prestanda, tillförlitlighet och anpassningsbarhet för produktionsmiljö.Nyttan med arbetet som presenteras i denna avhandling är att öka kunskapsbasen för hur pappers och flexibla filmers ytegenskaper påverkar prestandan på inkjet-tryckta nanopartikelstrukturer. Detta möjliggör bättre urval, anpassning av, eller tillverkning av sådana substrat för att passa tryckta mönster med hög konduktivitet; som till exempel tryckta antenner på förpackningar.

Serine proteases are a family of enzymes with a wide array of functions across both eukaryotes and prokaryotes. Here we have attempted to produce the serine proteases rat mast cell protease 2 and mouse mast cell protease 5 in a culture of HEK 293 cells; and mouse mast cell protease 4, platypus granzyme B-like protease and platypus hypothetical protease in a baculovirus expression system. Following production we wanted to analyse these serine proteases using a phage display assay and a battery of chromogenic substrates.

With the industry striving for smaller devices, new technologies are developed to further miniaturize electronics devices. To this end, realization of 3D/non-planar interconnects, which aim at miniaturizing the interconnects formed between components on the same device, has attracted many researchers. This thesis focuses on a feasibility analysis for developing non-planar interconnects on various flexible substrates using laser assisted maskless microdeposition (LAMM), which is a pressure-less process. There are two types of flexible substrates that are used: double-sided copper substrates separated by a layer of polyethylene terephthalate (PET) as well as a polyethylene terephthalate flexible substrate with surface-mounted resistors. For both substrates, multiple types of experiments were conducted to discover procedures which result in the highest rate of success for forming conductive interconnects. Optimal process parameters and deposition techniques were determined after multiple experiments. After experiments were completed, the resultant substrates were subject to various characterization methodologies including optical and scanning electron microscopy, energy-dispersive X-ray spectroscopy, X-ray diffraction and profilometery. The results of these methodologies are documented in this thesis. After many types of experiments involving substrate manipulation of the double-sided copper substrates, it was shown that the silver nano-particles were more likely to form a conductive interconnect when a polished slant was fabricated on the substrate. Many deposition patterns were used for the flexible substrates with surface-mounted resistors. Of these patterns, the two patterns, the ‘zigzag’ and ‘dot solder’ patterns, proved to have a much higher success rate for creating conductive interconnects compared to the other patterns. During this study, the results of the experiments using the LAMM process show that this technology has great potential for creating non-planar interconnects on flexible substrates. The experiments however suggest that the process is very sensitive to the material composition and process parameters. As such, with a small change in parameters, the 3D interconnects can fail to be produced. It was also observed that the possibility of silver interconnect fractures is higher where dissimilar materials with different thermal expansion rates are used for the underlying substrates.

Diamond-like Carbon (DLC) film exhibits an extreme hardness, low friction coefficient, chemical stability, heat conductivity, and high resistance. Their properties lead to remarkable applications on industry. In the experiment, we use electrondeposition to deposit the DLC film on Si substrate. Different concentrations of electrolyte were used to deposit on the of silicon substrates with different roughness surface. KOH solution was used to etch and to get the different roughness on the surface of silicon substrates. the morphology of surface were observed by SEM and AFM. Composition and microstructure of the DLC film were characterized by the Raman spectroscopy and XPS, repectively. The optical properties of DLC film were investigated by the N&K analyzer. From the AFM results, the surface morphology observed by KOH etching on the surface of silicon substrates during etching time as 0¡B20¡B40¡B60 min, the surface roughness increased from 2.64 to 14.07 nm. Based on thevariation of surface roughness, the growth rate was observed more quicker than the non etch surface. Moreover, to deposit the DLC film on the alkalinity solution was better then acid solution. However, the ID/IG ratio and the sp2/sp3 ratio obtained from Raman and XPS increase with the roughness surface from 1.09 to 1.82 and 0.985 to 2.15, respectively. It is because that the microstructure of DLC film varies and exchange to graphitization. The mixed the ammonia water and ammonium acetate into acetic acid solution was used to deposit DLC film on Si surface, and film shows with lower ID/IG ratio. Additionally, as the amount of ammonium acetate was varied in the solution, the ID/IG ratio of the films observed as decrease from 1.2 to 0.93 with increasing amount of ammonium acetate 10g to 40g. It was due to the methyl radicals increase in the solution. Besides, we can find the optical band gap decreased with DLC films changing to graphitization.

In the process of sputtering, what a system operator concerns are the sputtering rate, target utilization, and substrates uniformity. All of them are influenced by variables such as electromagnetic environment, chamber temperature, and pressure. In thin film manufacturing, targets bombarded by ions will sputter atoms to the substrates in order to make thin films; therefore, when a certain target zone is extensively bombarded by ions, target surface will become thinner. In general, when certain part of the target is penetrated, it is no longer usable while utilization rate only from 30 to 50 percent. It causes considerable waste and relatively higher costs. As a result, the objective of this study is to enhance target utilization and the sputtering rate through appropriate adjustment in the structure of the existing DC Magnetron Sputtering System (MSS). Since, the magnetic field distribution in the chamber will be appropriately adjusted inside the DC MSS with extra iron annulus and active compensation magnetizations being added. However, in order to get the better structural refinement of DC MSS it needs a thorough design and management based on Taguchi Method. Then, based on such structural adjustment, electron trajectories on top surface of targets can be conveniently controlled, and target erosion patterns and the number of ions bombarding the target will be indirectly controlled. It will, as a result, achieve the objective of this study by enhancing not only the target utilization efficiency but the sputtering rate.

Hydrogen, one of the cleanest energies, is very attractive in the near future. However, it could be hazardous to store, transport and use hydrogen gas because leakage can cause explosion if sparks appear. Therefore, it is essential to develop sensors to detect the hydrogen leakage in order to prevent potential accidents. In this research, Metal-Insulator-Semiconductor (MIS) Schottky-diode hydrogen sensors with different gate insulators (Ta2O5, La2O3, LaTiON, and HfTiO) or substrates (Si, SiC, and InGaN/GaN MQW) were prepared in order to study their hydrogen sensing performances. Firstly, two sensors based on Si and SiC with Ta2O5 as gate insulator were prepared and compared. Owing to high permittivity (~25), good thermal stability and low electrical defects, Ta2O5 was chosen as the insulator. The differences in sensitivity and response time between the two sensors were ascribed to the difference in the surface morphology of Ta2O5 between the SiC sensor (mean surface roughness was 0.39 nm) and its Si counterpart (mean surface roughness was 0.22 nm). Secondly, due to the high permittivity (~25) and good thermal stability of La2O3, the high permittivity (~20), low interface-state density, and low leakage current of LaTiON, Si sensors with these two dielectrics as gate insulator were developed. The sensitivity of the La2O3 sensor could exceed 7.0 at 150 oC, and the sensor exhibited good hydrogen sensing performance at up to 250 oC. On the other hand, the maximum sensitivity of the LaTiON sensor could reach 2.5 at 100 oC. For the LaTiON sensor, the Poole-Frenkel model controlled the carrier transport at high temperatures (150 ~ 200 oC) while the thermionic emission was the dominant conduction mechanism at lower temperatures (from room temperature to 150 oC). For the La2O3 sensor, the hydrogen reaction kinetics was confirmed, and an activation energy of 10.9 kcal/mol was obtained for this sensor. Thirdly, the La2O3 gate insulator used in the previous work was applied to make MIS sensor on SiC substrate for higher-temperature applications. Its maximum sensitivity and response time at high temperature (260 oC) are 4.6 and 20 s, respectively. The electrical conduction mechanisms were explained in terms of Fowler-Nordheim tunneling (below 120 oC) and the Poole-Frenkel effect (above 120 oC). Finally, in order to see whether the unique structure of InGaN/GaN multiple quantum wells (MQWs) can be utilized for the MIS Schottky-diode hydrogen sensor, three sensors were made on InGaN/GaN MQWs substrate, one without gate insulator, one Finally, in order to see whether the unique structure of InGaN/GaN multiple quantum wells (MQWs) can be utilized for the MIS Schottky-diode hydrogen sensor, three sensors were made on InGaN/GaN MQWs substrate, one without gate insulator, one In summary, the quality of the gate insulator plays an important part in the performance of the hydrogen sensors. SiC and InGaN/GaN MQW substrates are suitable for high-temperature (from ~200 to ~500 oC) applications while the low-cost sensors based on Si substrate can function well below about 200 oC. Hydrogen sensors with these high-k materials (Ta2O5, La2O3, LaTiON, and HfTiO) as gate insulator can produce good electrical characteristics, high sensitivity, and fast response. / published_or_final_version / Electrical and Electronic Engineering / Doctoral / Doctor of Philosophy

<p>The posttranslation modification of histone proteins within the nucleosomes of chromatin plays important roles in the regulation of gene expression in both normal biological and pathobiological processes. These modifications alter local chromatin structure and subsequently alter the expression profile of associated genes. Histone methylation, which was long thought immutable, is one such modification that plays a dual functionality in both activation and repression of gene expression and can be thought of as an information storage mark. With the initial discovery of lysine-specific demethylase 1A (LSD1/KDM1A), an FAD-dependent enzyme that catalyzes the oxidative demethylation of histone H3K4me1/2 and H3K9me1/2 repressing and activating transcription, respectively, the missing counterbalance to dynamic histone methylation was cemented. This discovery further strengthened the link between histone demethylation and transcriptional regulation and the enzyme has since been identified as a target with therapeutic potential.</p><p>Given the significance of KDM1A enzymatic activity, herein we report our efforts to characterize novel binding interactions that dictate the enzymes biological and pathobiological functions. As KDM1A falls into the greater class of flavin-dependent amine oxidases, it contains features that are recurrent within the class, but due to its unique ability to work on histone and non-histone substrates has unprecedented structural elements. Although the active site is expanded compared to the greater amine oxidase superfamily, it is too sterically restricted to encompass the minimal 21-mer peptide substrate footprint of the histone H3 tail. The remainder of the substrate/product is therefore expected to extend along the surface of KDM1A. Using steady-state kinetic analyses, we now show that unmodified histone H3 is a tight-binding, competitive inhibitor of KDM1A demethylation activity with a Ki of 18.9 ± 1.2 nM that is approximately 100-fold higher than the 21-mer peptide product. The relative affinity of dose-response curves is independent of preincubation time suggesting that H3 rapidly reaches equilibrium with KDM1A. Rapid dilution experiments confirmed the increased binding affinity of full-length H3 toward KDM1A was at least partially caused by a slow off-rate with a koff of 0.072 min-1, a half-life (t1/2) of 9.63 min, and residence time (τ) of 13.9 min. Independent affinity capture surface plasmon resonance experiments confirmed the tight-binding nature of the H3/KDM1A interaction revealing a Kd of 9.02 ± 2.27 nM, a kon of 9.26 x 104 ± 1.5 x 104 M-1s-1 and koff of 8.35 x 10-4 ± 3.4 x 105 s-1. Additionally, consistent with H3 being the only histone substrate of KDM1A, no other core histones are inhibitors of demethylation activity. Our data suggests that KDM1A contains a histone H3 secondary specificity recognition element on the enzyme surface and required further characterization.</p><p>In order to characterize this secondary H3 binding site, we turned to the use of cysteine labeling, chemical cross-linking coupled to proteolysis and LC-MS/MS, HDX-MS, and the design of an active, tower domain deletion KDM1A mutant. We now show that the tower domain contributes to the extended binding interface of the KDM1A/H3 interaction. Additionally, we show that the KDM1A tower domain is not required for demethylation activity and that one can functionally uncouple catalytic activity from protein-protein interactions that occur along the KDM1A tower domain interface, a domain unprecedented in the greater amine oxidase family. Furthermore, this towerless mutant will be useful for dissecting molecular contributions to KDM1A function along the tower domain. Our discovery of this secondary binding site within the aforementioned domain points to how pivotal this region is to the control and localization of KDM1A enzymatic activity as it also serves a pivotal role as a protein-protein interaction motif for the nucleation of a multitude of multimeric protein complexes.</p><p>With this in mind, we set out to design a strategy to isolate the core histone demethylase complex from E. coli cellular lysates. With the use of polycistronic vectors that encode both KDM1A and CoREST for coexpression we were able to produce appreciable amounts of chromatographically pure complex. As our CoREST construct in this strategy contains both the ELM2 and SANT2 domain needed for interaction with the HDACs, this core complex will serve as a starting point for future work that will tease apart additional influences on substrate binding and recognition imparted on KDM1A from binding partners. This preparation can therefore be used in a multitude of downstream studies including reconstitution of the core histone demethylase/deactylase complex and in depth kinetic and biophysical analyses and provides an invaluable starting point</p><p>This work provides a foundational understanding of this unprecedented secondary binding site on the surface of the KDM1A tower domain and how it may play an important role in substrate and product recognition. We suspect that this extended interaction interface may control KDM1A localization within specific chromatin loci and allow the enzyme to serve as a docking element for the nucleation of protein complexes or transcriptional machinery. On the other hand, disruption of this point of contact between the KDM1A/H3 binary complex may also facilitate enzyme/product dissociation, thereby tuning the catalytic activity of the demethylase. Additionally, the ability to produce substantial quantities of the core histone demethylase complex is a necessary step in the decoding of the ‘histone code’ hypothesis of KDM1A and its associated complexes. We suspect that the body of this work will prove to be invaluable for future characterization of the enzyme and its role in biology and pathobiology.</p> / Dissertation