Applications such as satellite and digital mobile radio systems (DMR) have gained widespread acceptance in recent years, and efficient digital processing techniques are gradually replacing the older analogue systems. An important subsystem of these applications is voiceband communication, especially digital speech encoding. Digital encoding of speech has been a focus of speech processing research for many years, and recently this activity together with the rapid advances in digital hardware, has begun to produce realistic working algorithms. This is typified by the Pan-European DMR system which operates at 13Kbit/s. For applications operating below this coding capacity, sophisticated algorithms have been developed. A particular class of these, termed Analysis-by-Synthesis Linear Predictive Coding (ABS-LPC), has been a subject of active world-wide research. In this thesis, ABS-LPC algorithms are investigated with particular emphasis on the Code-Excited Linear Predictive coding (CELP) variant. The aim of the research is to produce high communication quality speech at 8Kbit/s and below by considering aspects of quantisation, computational complexity and robustness. The ABS-LPC algorithms operate by exploiting short-term and long-term correlations of speech signals. Line Spectral Frequency (LSF) representation of the short-term correlation is examined and various LSF derivations and quantisation procedures are presented. The variants of ABS-LPC are compared for their advantages and disadvantages to determine an algorithm suitable for in-depth analysis. The particular chosen variant, CELP, was pursued. A study on the importance of the long-term prediction, and the simplification of CELP without sacrificing speech quality is presented. The derived alternative approaches for the computation of the long-term predictor and the filter excitation have enabled the previously unpractical CELP algorithm to produce high communication quality speech at rates below 8Kbit/s, and yet remain implement able in real-time on a single chip. Refinements of the CELP algorithm followed in order to improve the coder towards higher speech quality at 4.8Kbit/s and below. This involved the examination of the weaknesses of the basic CELP algorithm, and alternative strategies to overcome these limitations are presented.

No description available.

Phase key, which uses optical encoding techniques for system security, plays an important role in optical storage, optical communication, and optical display. It employs a random phase generator with a volume hologram for optical encoding. The advantages of using phase keys for optical communication is: (1) it is hard to be duplicated and (2) it requires sensitively alignment to decode the desired signal. Thus, it ensures security of the optical system. However, the adjunctive challenge of using a phase key is the difficulty of alignment by users. In this paper, we propose a method for optical alignment of the phase key in a random phase encoded volume holographic storage system. In this method, a holographic correlator is applied to help the optical alignment of the phase key. It has been shown that the desired signal from the random phase encoded volume holographic storage system can be retrieved easily with high security.

<p>    </p> <p>DNA-encoded chemical libraries (DELs) are collections of DNA-linked small molecules, where each synthetic small molecule is covalently attached to a unique DNA barcode that encodes its identity. This technology harnesses the power of organic chemistry and genetics, which extends the application of molecular evolution and natural selection to the discovery of specific small molecules binders to protein targets of interest. Rather than discretely screening individual molecules, up to billions of DNA-encoded small molecules can be assessed collectively by a selection assay in a single tube. As a result, the high sensitivity, low cost, and unprecedented level of molecular complexity of DELs allow rapid generation of novel bioactive compounds. While powerful, this approach has its own limitations, including limited target scope and selection strategies. Currently, DEL targets have been largely limited to biochemically purified proteins and used in affinity-based selections assays. In the first area of this work, we address both these limitations by capitalizing on the power of affinity labeling. This allows DELs to be applied to protein targets within and on living cells and expands the power of DNA-encoding to the identification of small molecules with specific biological functions beyond binding. </p> <p>In the second area, we harnessed affinity labeling and DNA sequence analysis to develop multiplexed small molecule ligand binding assays. This method is the initial demonstration of split-and-pool ligand binding assays using DNA-linked small molecule probes. We used this approach in a high-throughput screening campaign to identify selective inhibitors by screening 1000 compounds against 5 bromodomain proteins concurrently. In addition, this approach was utilized to rank order the affinity of a 96-member library of DNA-linked ligands to a protein simultaneously, which significantly increases the throughput of ligand binding assays while keeps the cost low. </p> <p>Lastly, we developed proximity-induced selection assays to enrich ligands from DELs. This approach involves uncaging or installation of a biotin purification tag on the DNA construct either through photo-deprotection of a protected biotin group using a light emitting protein tag or by amine acylation using an engineered biotin ligase. Compared to affinity labeling-based selection approaches, this approach results in improved recovery of ligands and, at the same time, removes the onerous requirement of protein purification. The enzyme-mediated proximity labeling approach should serve as a convenient tool for molecular discovery with DELs. </p>

MA, School of Social Sciences, Faculty of Humanities, University of the Witwatersrand, 2002

A thesis submitted to the Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, in fulfilment of the requirements for the degree of Doctor of Philosophy Johannesburg, 2002 / Hepatitis B virus (HBV) infection is endemic to several populous regions and is often complicated by cirrhosis and hepatocellular carcinoma (HCC). Present treatment of chronic HBV infection is usually ineffective and novel therapeutic approaches are an important medical objective. The X open reading frame (ORF) of HBV, HBx, is a conserved sequence that overlaps with the polymerase ORF and viral c/'s-elements, and is present within all viral transcripts. In addition, the HBx ORF encodes a 17 kDa transactivator protein, HBx, which is required for the establishment of viral infection and has been implicated in HBV-associated hepatocarcinogenesis. The HBx sequence thus represents a compelling target for applying nucleic acid hybridisation-based therapeutic agents for the inhibition of HBV gene expression and replication. / IT2018

There are many diverse classes of biologically active natural products containing chiral 1,5- or 1,5,7-polyol moieties, including the novel fibrinogen receptor antagonist tetrafibricin, a potential antiplatelet therapeutic drug to treat various arterial thrombotic diseases. There have been some elegant synthetic strategies developed to synthesize these challenging 1,5-diol motifs; however, many of them suffer from a variety of inherent limitations. To overcome many of these challenges, our group has developed an iterative configuration-encoded strategy to access 1,5-polyols with unambiguous stereocontrol by exploiting Julia–Kocienski couplings of enantiopure α-siloxy-γ-sulfononitrile building blocks with alcohol stereocenters previously established via asymmetric catalysis. Our method is a strategy level innovation that allows for the efficient and rapid access to all stereoisomers of a 1,5-polyol family from cheap and easily accessible reagents, without the need to determine the configuration of each alcohol stereocenter in the growing polyol chain. We were able to modify our configuration-encoded 1,5-polyol methodology to access the anti,syn-1,5,7-triol within the C15–C25 fragment of tetrafibricin with excellent selectivity by incorporating differentiable protection and merging this approach with the tactic of diastereoselective intramolecular conjugate addition via benzylidene acetal construction to access the syn-1,3-diol functionality. We also applied our iterative configuration-encoded strategy to the synthesis of the 1,5-polyol-containing C26–C40 fragment of tetrafibricin with excellent stereoselectivity by modifying our previous route to the C27–C40 segment. By overcoming the challenges associated with the reduction of α-siloxynitriles and extending the carbon chain to alter the subsequent Mukaiyama aldol coupling location, we were able to furnish the C26–C40 fragment with the correct protection and functionality for further coupling to the C15–C25 segment. With the C15–C25 and C26–C40 fragments in hand, we joined these segments via asymmetric BF3⋅OEt2-mediated Mukaiyama aldol construction with high 1,3-anti stereoinduction. We determined the preceding stereoselectivity by first using the simplified model C26–C40 fragment and found that replacing the TBDPS with TBS protection of the β-siloxy aldehyde increased the level of 1,3-anti induction. To complete the C15–C40 fragment of tetrafibricin, we performed an intramolecular hydroxyl-directed anti-reduction to furnish the desired anti,anti,anti-1,3,5,7-tetraol moiety. We were able to establish the configurations of these chiral alcohols using a battery of 2D NMR experiments. Finally, to complete the total synthesis of tetrafibricin, we have proposed a route to couple our C15–C40 fragment with the C8–C14 segment via a precedented asymmetric aldol reaction, followed by coupling to the known C1–C7 polyene fragment. With minor functional group transformations and a global deprotection, access to the natural product tetrafibricin should be achievable.

In 2008 the Nobel Prize in Chemistry was awarded to the scientists who revolutionized biomedical technology by isolating, characterizing, and pioneering the use of a green fluorescent protein (GFP) from a humble hydrozoan jellyfish. Now numbering in the hundreds of colors and applications, fluorescent protein (FP) tools have facilitated the explosion of biological knowledge elucidated by a technology that can label DNA or RNA, track protein expression, and identify protein interactions. The development of the large variety of FP biotechnology available today has been due to the need for expanded color palettes and applications, and more efficient functionality. Yet, as our understanding of the biochemical and spectral characteristics of these genetically-encoded, self-assembling proteins has expanded, our comprehension of the biological function of FPs in the host organisms has remained inadequate. While the need for novel FP laboratory applications still continues, the new focus in the field of fluorescent proteins is moving to also characterize their biological functions. In this research compilation, the identification of three groups of new fluorescent proteins from marine copepods and hydrozoans has provided a collection of eleven FPs exhibiting previously uncharacterized colors, and biochemical and structural features. The green FPs from copepods are the brightest wild-type FPs identified and support the hypothesized biological function of fluorescence as counter-shading in the marine environment where these animals live. The FPs from the siphonophore and anthoathecate jelly, both hydrozoan animals, are comprised of tandemly expressed fluorescent protein units, a solution to the oligomeric structure common to most FPs that suggests a novel structure-function relationship. The fluorescent proteins from Obelia reveal a novel hydrozoan cyan FP, previously uncharacterized higher-order structural complexes, and have initiated the work to describe the biological function of these proteins as potential regenerators of their internal bioluminescent light sources. All eleven fluorescent proteins may also be adapted for FP technology. / text

Advances in genome and proteome research have led to a dramatic increase in the number of macromolecular targets of interest to the life sciences. A solution-phase method to simultaneously reveal all ligand-target binding pairs from a single solution containing libraries of ligands and targets could significantly increase the efficiency and effectiveness of target-oriented screening efforts. Here, we describe interaction-dependent PCR (IDPCR), a solution-phase method to identify binding partners from combined libraries of small-molecule ligands and targets in a single experiment. Binding between DNA-linked targets and DNA-linked ligands induces formation of an extendable duplex. Extension links codes identifying the ligand and target into one selectively amplifiable DNA molecule. In a model selection, IDPCR resulted in the enrichment of DNA encoding all five known protein-ligand pairs out of 67,599 possible sequences.

The spatio-temporal attributes of intracellular calcium (Ca2+) transients activate various biological functions. These Ca2+ signaling events are triggered extracellularly through different stimuli and controlled intracellularly by the major Ca2+ storage organelle and by numerous Ca2+ pumps, channels, and Ca2+ binding proteins. Ca2+ transients can be significantly altered as a result of defects with signal modulation, leading to different diseases. Because of the fragility and intricacy of the Ca2+ signaling system, with the endo- and sarcoplasmic reticulum at the center, genetically-encoded Ca2+ probes that have been optimized for mammalian expression and fast kinetics are needed to observe global and local Ca2+ changes in different cells. Here, we first report the crystal structure determination of our genetically-encoded Ca2+ sensor CatchER which utilizes EGFP as the scaffold protein. Crystal structures of CatchER were resolved in the Ca2+-free, Ca2+-loaded, and gadolinium-loaded forms at 1.66, 1.20, and 1.78 Å, respectively. Analysis of all three structures established conformational changes in T203 and E222 produce the varying ratios of the neutral and anionic chromophore reflected in the absorbance spectrum where Ca2+ stabilizes the anionic chromophore and enhances the optical output. Since CatchER has miniscule fluorescence when expressed at 37˚C in mammalian cells, we enhanced its brightness by improving the folding at 37˚C, facilitating better chromophore formation. The resulting mutants are the CatchER-T series of Ca2+ sensors with CatchER-T’ having the most improvement in brightness at 37˚C. We also introduced the N149E mutation in the binding site to alter the Kd along with the brightness mutations. The resulting mutants were characterized and found to have weaker Kds compared to wild-type CatchER, similar quantum yields, and altered ratios of the neutral and anionic chromophore in the apo form. Then, CatchER-T’ was applied in situ to monitor Ca2+ changes globally in the ER/SR of C2C12, HEK293, and Cos-7 cells. A new construct consisting of CatchER-T’ and JP-45 was created to monitor local Ca2+ dynamics in the SR lumen of skeletal muscle cells. The results showed a difference between global and local SR Ca2+ release. We also examined the potential and spectroscopic properties to utilize some of our sensors in T cells to monitor the magnesium (Mg2+) flux in immune cells with faulty MagT1 receptors to understand the role of Mg2+ in the immune response.