The intent of this research is to create a qualitative framework to guide perceptions and observations about informal self-directed learning in the arena of contemporary Web 2.0 e-learning. Drawing influence from the studies on self-directed learning conducted by Allen Tough and his associates, the writings on educational connoisseurship and criticism by Elliot Eisner, as well as research and literature about contemporary e-learning contexts, this exploratory study is comprised of a hermeneutic analysis that seeks to discover themes, patterns and points of intersection in these three areas. The analysis presents the application of the resulting framework to three illustrative scenarios, constructed from ideas and themes drawn from the major elements of the study, and discusses the findings revealed by the investigation. The study concludes with reflection and recommendations for application and further research.

<p><i>Lactococcus lactis,</i> prolidase (<i>Lla</i>prol) hydrolyzes Xaa-Pro dipeptides. Since Xaa-Pro is known as bitter peptides, <i>Lla</i>prol is potentially applicable to reduce bitterness of fermented foods. <i>Lla</i>prol shows allosteric behaviour and substrate inhibition, which are not reported in other prolidases. Computer models of <i>Lla</i>prol based on an X-ray structure of non-allosteric <i>Pyrococcus furiosus</i> prolidase showed that a loop structure (Loop^32-43) is located at the interface of the protomers of this homodimeric metallodipeptidase. This study investigated roles of four charged residues (Asp³⁶, His³⁸, Glu³⁹, and Arg⁴⁰) of Loop^32-43 in <i>Lla</i>prol using a combination of kinetic examinations of ten mutant enzymes and their molecular models. Deletion of the loop structure by Î36-40 mutant resulted in a loss of activity, indicating Loop^32-43 is crucial for the activity of <i>Lla</i>prol. D36S and H38S exhibited 96.2 % and 10.3 % activity of WT, whereas little activities (less than 1.0 % of WT activity) were observed for mutants E39S, D36S/E39S, R40S, R40E, R40K and H38S/R40S. These results implied that Glu³⁹ and/or Arg⁴⁰ play critical role(s) in maintaining the catalytic activity of <i>Lla</i>prol. These observations suggested that the loop structure is flexible and this attribute, relying on charge-charge interactions contributed by Arg⁴⁰, Glu³⁹ and Lys¹⁰⁸, is important in maintaining the activity of <i>Lla</i>prol. When the loop takes a conformation close to the active site (closed state), Asp³⁶ and His³⁸ at the tip of the loop can be involved in the catalytic reaction of <i>Lla</i>prol. The two active mutant prolidases (D36S and H38S) resulted in modifications of the unique characteristics; the allosteric behaviour was not observed for D36S, and H38S <i>Lla</i>prol showed no substrate inhibition. D36E/R293K, maintaining the negative charge of position 36 and positive charge of position 293, still possessed the allosteric behaviour, whereas the loss of the charges at these positions (D36S of this study and R293S of a previous study (Zhang et al., 2009 BBA-Proteins Proteom 1794, 968-975) eliminated the allosteric behaviour. These results indicated the charge-charge attraction between Asp³⁶ and Arg²⁹³ is important for the allostery of <i>Lla</i>prol. In the presence of either zinc or manganese divalent cations as the metal catalytic centre, D36S and H38S enzymes also showed different substrate preferences from WT <i>Lla</i>prol, implying the influence of Asp³⁶ and His³⁸ on the substrate binding. D36S and H38S also showed higher activities at pH 5.0 to 6.0, in which range WT <i>Lla</i>prol steeply decreased its activity, indicating Asp³⁶ and His³⁸ are involved in the active centre and influence the microenvironment of catalytic His²⁹⁶. The above observations are attributed to modifications of their local structure in the active centre since the temperature dependency and thermal denaturing temperature indicated little effects on the overall structure of the <i>Lla</i>prol mutants.</p> <p>From these results, we concluded that the unique behaviours of <i>Lla</i>prol are correlated to Loop^32-43 and Asp³⁶ and His³⁸ on it. When Loop^32-43 takes a closed conformation, Asp³⁶ interacts with Arg²⁹³ via charge-charge attraction to form an allosteric subsite. The saturation of the allosteric site with substrates further allowed the communications of His³⁸ with S₁ site residues to complete the active site. When the substrate concentration becomes higher than it is required to saturated productive S₁' site, His³⁸, Phe¹⁹⁰ and Arg²⁹³ would resemble the residue arrangement of S₁' site residues (His²⁹², Tyr³²⁹, and Arg³³⁷) and bind to the proline residue of substrates. This non-productive binding would prevent the conformational change of Loop^32-43, which further results in the substrate inhibition. For further confirmation of this mechanism, crystallographic studies will be conducted. In this thesis, we have indentified the conditions to produce crystals of <i>Lla</i>prol proteins.</p>

Haskell is a lazy functional language with a strong static type system and excellent support for parallel programming. The language features of Haskell make it easier to write correct and maintainable programs, but execution speed often suffers from the high levels of abstraction. While much past research focuses on high-level optimizations that take advantage of the functional properties of Haskell, relatively little attention has been paid to the optimization opportunities in the low-level imperative code generated during translation to machine code. One problem with current low-level optimizations is that their effectiveness is limited by the obscured control flow caused by Haskell's high-level abstractions. My thesis is that trace-based optimization techniques can be used to improve the effectiveness of low-level optimizations for Haskell programs. I claim three unique contributions in this work. The first contribution is to expose some properties of low-level Haskell codes by looking at the mix of operations performed by the selected benchmark codes and comparing them to the low-level codes coming from traditional programming languages. The low-level measurements reveal that the control flow is obscured by indirect jumps caused by the implementation of lazy evaluation, higher-order functions, and the separately managed stacks used by Haskell programs. My second contribution is a study on the effectiveness of a dynamic binary trace-based optimizer running on Haskell programs. My results show that while viable program traces frequently occur in Haskell programs the overhead associated with maintaing the traces in a dynamic optimization system outweigh the benefits we get from running the traces. To reduce the runtime overheads, I explore a way to find traces in a separate profiling step. My final contribution is to build and evaluate a static trace-based optimizer for Haskell programs. The static optimizer uses profiling data to find traces in a Haskell program and then restructures the code around the traces to increase the scope available to the low-level optimizer. My results show that we can successfully build traces in Haskell programs, and the optimized code yields a speedup over existing low-level optimizers of up to 86% with an average speedup of 5% across 32 benchmarks.

Progress in DNA sequencing of plant genomes has revealed that, in addition to microorganisms, a number of plants contain genes which share similarity to microbial 1-aminocyclopropane-1-carboxylate (ACC) deaminases. ACC deaminases break down ACC, the immediate precursor of ethylene in plants, into ammonia and α-ketobutyrate. We therefore sought to isolate putative ACC deaminase cDNAs from tomato plants with the objective of establishing whether the product of this gene is a functional ACC deaminase. It was demonstrated that the enzyme encoded by the putative ACC deaminase cDNA does not have the ability to break the cyclopropane ring of ACC, but rather that it utilizes D-cysteine as a substrate, and in fact encodes a D-cysteine desulfhydrase. Kinetic characterization of the enzyme has shown that it is similar to other previously characterized D-cysteine desulfhydrases. Using site-directed mutagenesis, it was shown that altering two amino acid residues within the predicted active site changed the enzyme from D-cysteine desulfhydrase to ACC deaminase. Concomitantly, it was shown that by altering two amino acids residues at the same position within the active site of ACC deaminase from Pseudomonas putida UW4 changed this enzyme into D-cysteine desulfhydrase.

Search-based placement methods have long been used for placing integrated circuits targeting the field programmable gate array (FPGA) and standard cell design styles. Such methods offer the potential for high-quality solutions but often come at the cost of long run-times compared to alternative methods. This dissertation examines strategies for enhancing local search heuristics---and in particular, simulated annealing---through the application of directed moves. These moves help to guide a search-based optimizer by focusing efforts on states which are most likely to yield productive improvement, effectively pruning the size of the search space. The engineering theory and implementation details of directed moves are discussed in the context of both field programmable gate array and standard cell designs. This work explores the ways in which such moves can be used to improve the quality of FPGA placements, improve the robustness of floorplan repair and legalization methods for mixed-size standard cell designs, and enhance the quality of detailed placement for standard cell circuits. The analysis presented herein confirms the validity and efficacy of directed moves, and supports the use of such heuristics within various optimization frameworks.

<p><i>Lactococcus lactis,</i> prolidase (<i>Lla</i>prol) hydrolyzes Xaa-Pro dipeptides. Since Xaa-Pro is known as bitter peptides, <i>Lla</i>prol is potentially applicable to reduce bitterness of fermented foods. <i>Lla</i>prol shows allosteric behaviour and substrate inhibition, which are not reported in other prolidases. Computer models of <i>Lla</i>prol based on an X-ray structure of non-allosteric <i>Pyrococcus furiosus</i> prolidase showed that a loop structure (Loop^32-43) is located at the interface of the protomers of this homodimeric metallodipeptidase. This study investigated roles of four charged residues (Asp³⁶, His³⁸, Glu³⁹, and Arg⁴⁰) of Loop^32-43 in <i>Lla</i>prol using a combination of kinetic examinations of ten mutant enzymes and their molecular models. Deletion of the loop structure by Î36-40 mutant resulted in a loss of activity, indicating Loop^32-43 is crucial for the activity of <i>Lla</i>prol. D36S and H38S exhibited 96.2 % and 10.3 % activity of WT, whereas little activities (less than 1.0 % of WT activity) were observed for mutants E39S, D36S/E39S, R40S, R40E, R40K and H38S/R40S. These results implied that Glu³⁹ and/or Arg⁴⁰ play critical role(s) in maintaining the catalytic activity of <i>Lla</i>prol. These observations suggested that the loop structure is flexible and this attribute, relying on charge-charge interactions contributed by Arg⁴⁰, Glu³⁹ and Lys¹⁰⁸, is important in maintaining the activity of <i>Lla</i>prol. When the loop takes a conformation close to the active site (closed state), Asp³⁶ and His³⁸ at the tip of the loop can be involved in the catalytic reaction of <i>Lla</i>prol. The two active mutant prolidases (D36S and H38S) resulted in modifications of the unique characteristics; the allosteric behaviour was not observed for D36S, and H38S <i>Lla</i>prol showed no substrate inhibition. D36E/R293K, maintaining the negative charge of position 36 and positive charge of position 293, still possessed the allosteric behaviour, whereas the loss of the charges at these positions (D36S of this study and R293S of a previous study (Zhang et al., 2009 BBA-Proteins Proteom 1794, 968-975) eliminated the allosteric behaviour. These results indicated the charge-charge attraction between Asp³⁶ and Arg²⁹³ is important for the allostery of <i>Lla</i>prol. In the presence of either zinc or manganese divalent cations as the metal catalytic centre, D36S and H38S enzymes also showed different substrate preferences from WT <i>Lla</i>prol, implying the influence of Asp³⁶ and His³⁸ on the substrate binding. D36S and H38S also showed higher activities at pH 5.0 to 6.0, in which range WT <i>Lla</i>prol steeply decreased its activity, indicating Asp³⁶ and His³⁸ are involved in the active centre and influence the microenvironment of catalytic His²⁹⁶. The above observations are attributed to modifications of their local structure in the active centre since the temperature dependency and thermal denaturing temperature indicated little effects on the overall structure of the <i>Lla</i>prol mutants.</p> <p>From these results, we concluded that the unique behaviours of <i>Lla</i>prol are correlated to Loop^32-43 and Asp³⁶ and His³⁸ on it. When Loop^32-43 takes a closed conformation, Asp³⁶ interacts with Arg²⁹³ via charge-charge attraction to form an allosteric subsite. The saturation of the allosteric site with substrates further allowed the communications of His³⁸ with S₁ site residues to complete the active site. When the substrate concentration becomes higher than it is required to saturated productive S₁' site, His³⁸, Phe¹⁹⁰ and Arg²⁹³ would resemble the residue arrangement of S₁' site residues (His²⁹², Tyr³²⁹, and Arg³³⁷) and bind to the proline residue of substrates. This non-productive binding would prevent the conformational change of Loop^32-43, which further results in the substrate inhibition. For further confirmation of this mechanism, crystallographic studies will be conducted. In this thesis, we have indentified the conditions to produce crystals of <i>Lla</i>prol proteins.</p>

Progress in DNA sequencing of plant genomes has revealed that, in addition to microorganisms, a number of plants contain genes which share similarity to microbial 1-aminocyclopropane-1-carboxylate (ACC) deaminases. ACC deaminases break down ACC, the immediate precursor of ethylene in plants, into ammonia and α-ketobutyrate. We therefore sought to isolate putative ACC deaminase cDNAs from tomato plants with the objective of establishing whether the product of this gene is a functional ACC deaminase. It was demonstrated that the enzyme encoded by the putative ACC deaminase cDNA does not have the ability to break the cyclopropane ring of ACC, but rather that it utilizes D-cysteine as a substrate, and in fact encodes a D-cysteine desulfhydrase. Kinetic characterization of the enzyme has shown that it is similar to other previously characterized D-cysteine desulfhydrases. Using site-directed mutagenesis, it was shown that altering two amino acid residues within the predicted active site changed the enzyme from D-cysteine desulfhydrase to ACC deaminase. Concomitantly, it was shown that by altering two amino acids residues at the same position within the active site of ACC deaminase from Pseudomonas putida UW4 changed this enzyme into D-cysteine desulfhydrase.

Search-based placement methods have long been used for placing integrated circuits targeting the field programmable gate array (FPGA) and standard cell design styles. Such methods offer the potential for high-quality solutions but often come at the cost of long run-times compared to alternative methods. This dissertation examines strategies for enhancing local search heuristics---and in particular, simulated annealing---through the application of directed moves. These moves help to guide a search-based optimizer by focusing efforts on states which are most likely to yield productive improvement, effectively pruning the size of the search space. The engineering theory and implementation details of directed moves are discussed in the context of both field programmable gate array and standard cell designs. This work explores the ways in which such moves can be used to improve the quality of FPGA placements, improve the robustness of floorplan repair and legalization methods for mixed-size standard cell designs, and enhance the quality of detailed placement for standard cell circuits. The analysis presented herein confirms the validity and efficacy of directed moves, and supports the use of such heuristics within various optimization frameworks.

We study a directed network design problem called the $k$-$(S,T)$-connectivity problem; we design and analyze approximation algorithms and give hardness results. For each positive integer $k$, the minimum cost $k$-vertex connected spanning subgraph problem is a special case of the $k$-$(S,T)$-connectivity problem. We defer precise statements of the problem and of our results to the introduction. For $k=1$, we call the problem the $(S,T)$-connectivity problem. We study three variants of the problem: the standard $(S,T)$-connectivity problem, the relaxed $(S,T)$-connectivity problem, and the unrestricted $(S,T)$-connectivity problem. We give hardness results for these three variants. We design a $2$-approximation algorithm for the standard $(S,T)$-connectivity problem. We design tight approximation algorithms for the relaxed $(S,T)$-connectivity problem and one of its special cases. For any $k$, we give an $O(\log k\log n)$-approximation algorithm, where $n$ denotes the number of vertices. The approximation guarantee almost matches the best approximation guarantee known for the minimum cost $k$-vertex connected spanning subgraph problem which is $O(\log k\log\frac{n}{n-k})$ due to Nutov in 2009.

The intent of this research is to create a qualitative framework to guide perceptions and observations about informal self-directed learning in the arena of contemporary Web 2.0 e-learning. Drawing influence from the studies on self-directed learning conducted by Allen Tough and his associates, the writings on educational connoisseurship and criticism by Elliot Eisner, as well as research and literature about contemporary e-learning contexts, this exploratory study is comprised of a hermeneutic analysis that seeks to discover themes, patterns and points of intersection in these three areas. The analysis presents the application of the resulting framework to three illustrative scenarios, constructed from ideas and themes drawn from the major elements of the study, and discusses the findings revealed by the investigation. The study concludes with reflection and recommendations for application and further research.