The main purpose of this study was to compare the effectiveness of the cooperative learning based on conceptual change conditions and traditionally designed science instruction on 7th grade students&rsquo / understanding of chemical and physical changes and classification of matter concepts and attitudes toward science as a school subject. In this study 102 seventh grade students from four classes of a Science Course instructed by the two teachers from ODT&Uuml / G.V. &Ouml / zel ilk&ouml / gretim Okulu took part. One of the classes of each teacher was randomly assigned as experimental group, which were instructed with cooperative learning based on conceptual change conditions and the other classes were assigned as control group, which were instructed traditionally. This study was conducted during the 2004-2005 fall semester over a period of four weeks. In this study, to examine the effect of the treatment on dependent variables / science achievement related to chemical and physical changes and classification of matter concepts measured with Classification and Changes of Matter Concepts Test, and science attitude scores measured with Attitude Scale Toward Science as a school subject. Science Process Skills Test was used at the beginning of the study to determine students&rsquo / science process skills. ANCOVA and ANOVA were used testing the hypotheses of the study. The results showed that the cooperative learning based on conceptual change conditions group had a significantly higher scores with respect to achievement related to chemical and physical changes and classification of matter concepts than the traditionally designed science instruction group. However, there is no significant difference between the mean scores of cooperative learning based on conceptual change conditions group and traditionally designed science instruction group with respect to attitudes toward science as a school subject. Science process skills were a strong predictor for the achievement related to chemical and physical changes and classification of matter concepts. It may be useful to use the results of this study and instruments and strategies developed for this study for classroom teachers in order to help students to reduce or eliminate their misconceptions.

Secondary hardening ultra-high strength (SHUHS) steels possess a unique combination of strength, fracture toughness and stress corrosion cracking resistance, which makes them candidate materials for aircraft landing gear and armour applications. There is a sustained drive to develop stronger and tougher materials for such applications. The objectives of this thesis are two-fold: first, to develop a new SHUHS alloy that is stronger than the existing SHUHS steel developed at Defence Metallurgical Research Laboratory (DMRL), Hyderabad and second, to establish processing-structure-property correlations for the new alloy. Empirical design and development of these complex steels involves enormous effort, cost, time and materials resources. To avoid this, a semi-empirical approach was espoused in this thesis wherein thermodynamic calculations using ThermoCalc were conducted to computationally design a series of alloys with varying levels of Cr and Mo. The design space was constrained by two objectives related to M2C carbides which are the primary cause of secondary hardening in these alloys. The first objective was to increase the amount of M2C to increase the peak strength, while the second objective was to lower the Cr/Mo ratio of the M2C to control its over-ageing behavior. Two new alloys C23 (with 2Cr-3Mo, wt. %) and C55 (with 5Cr-5Mo, wt. %) and a base alloy akin to the DMRL SHUHS steel, C21 were selected for experimental validation. These alloys were melted, rolled and subjected to a battery of heat treatments. Austenitization studies revealed that the new alloys required higher austenitization temperatures to dissolve primary carbides. However such a treatment also resulted in an austenite composition that was not conducive for obtaining a fully martensitic microstructure on quenching. Based on these studies, the design space was modified to include additional criteria related to the Ms and precipitate dissolution temperatures. C55 failed to clear either criteria, while C23 cleared both, and so tempering studies were limited to C23. Isochronal tempering studies revealed that C23 in the peak aged condition was >10% stronger than C21 indicating that the alloy design objective of strength enhancement was achieved successfully. Microstructural characterization revealed that the strength enhancement was due to the higher number density and volume fraction of the M2C-like solute clusters in C23, which resist shearing in the under-aged condition and strengthen by Orowan mechanism in the over-aged condition. This thesis has successfully demonstrated that the design paradigm of enhancing strength by increasing the amount of M2C is justified and that ThermoCalc can be used to as an objective-oriented alloy design tool in this class of the steels.

One of the base intention of this research in didactic sphere is to describe and to analyse curriculum in all of its forms. In educational sphere focused on Information and Communication Technology (ICT) or on Informatics, this intention is very actual mainly due to recent integration to compulsory part of curriculum at elementary schools and also due to its dynamic development. This PhD thesis follows researches analysing curriculum of other educational spheres and also follows survey devoted to implementation of ICT to educational environment. Second type of research uses mainly quantitative method and relied on declarations of respondents. Compared to this, there was chosen qualitative case study getting data also from analysis of documents or from observing. At first, there are identified terms of curriculum, its phases and ICT literacy which presents one of the main goals of educational environment in the Czech curriculum. Then it is fol-lowed by analysis and summary of results of already realized surveys focused on teaching conditions of ICT and with ICT, together with informatics themes and their relations to individual actors. Last but not least, it is finished by results of students in ICT literacy and informatics thinking. Based on analysis of documents, the empirical part of theses describes models used to definition of educational sphere of ICT and projected curriculum of chosen basic school. Realized curriculum and its changes are deduced from analysis of notes in class-book. Videorecords of some lessons were used to verification of notes and also to analyse used educational methods. Based on its analysis, there were defined also educational goals of these teaching units and reached curriculum. Special practical exercise was used to verify the knowledge and skills of selected knowledge and skills. This thesis describes not only individual phases of ICT curriculum in chosen basic school but also identifies some differences between them. Finally, some of findings are used also as incentives to adjust the projected curriculum at the national level.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / In many applications of metalworking industry it is required mass production of parts containing holes with finish and tolerances typically from precision assembling, with working quality, ISO IT grade, ranging from IT7- IT5. But to produce these holes with tighter tolerances and with low geometrical errors, conventional twist drills may be not enough, so further processes are required, like reaming for example. Before any reaming process comes the drilling process, resulting in two different tools, two parts on necessary machining time, which in turn affects productivity and competitiveness. Therefore , any investment in the production of tools and techniques to reduce the machining time of close tolerance holes without affecting the integrity of the surface and preferably at a lower cost , will receive special attention from the machining community. In this context, this paper presents a technical and economic study of the use of special designed drill in replacement by a reamer the production of cylindrical holes of 10 mm diameter pieces of compacted graphite cast iron in terms of finish, roundness and dimensional deviation. It was also investigated the influence of the allowance in the final quality of the machined hole reamer. The surface of the holes and the microhardness of the machined holes were also analyzed and the tool wear was monitored. Finally, the economic analysis of the tools employed was performed. The results showed that special drill resulted in finish and dimensional deviation as good as those produced by the reamer. IT6 was the working quality obtained after reaming with reaming allowance of 0.5mm, while IT8 quality was obtained by using the special designed drill. Wear on special drill was lower than that observed in the reamer, and for the latter chipping occurred. Up to 29% of reduction in machining time was achieved by using the special drill, which resulted in a reduction of 33% cost as well. In contrast the values of cylindricity deviations were higher than those obtained with the reamers under the conditions investigated. The hardness was not affected by the processes employed. / Em muitas aplicações da indústria metal-mecânica exige-se a fabricação de peças com furos que apresentem acabamento e exatidão geométrica necessária para a montagem de precisão, normalmente com qualidade de trabalho variando entre ISO IT5-IT7. Mas para a produção destes furos de tolerâncias mais estreitas e com menores erros geométricos, as brocas helicoidais convencionais não são suficientes, de forma que são necessários processos subsequentes, como o alargamento, por exemplo. Antes de qualquer operação de alargamento vem à operação de furação, chamada de pré-furação, e assim para um furo alargado são necessárias no mínimo duas ferramentas diferentes, duas parcelas no tempo de usinagem, que por sua vez afeta a produtividade e competitividade. Portanto, qualquer investimento na produção de ferramentas e técnicas que permitam reduzir o tempo de usinagem de furos de tolerância estreita, sem afetar a integridade da superfície e de preferência com menor custo, receberá especial atenção da comunidade de usinagem. Neste contexto, este trabalho apresenta um estudo técnico e econômico do emprego de broca especial em substituição ao alargador, na produção de furos cilíndricos, de 10 mm de diâmetro, em peças de ferro fundido vermicular, em termos de acabamento, cilindricidade e desvio dimensional. Investigou-se também a influência do sobremetal na qualidade final do furo usinado pelo alargador. As paredes dos furos e a microdureza dos furos usinados foram também analisados e o desgaste das ferramentas foi monitorado. Por último, foi realizada a análise econômica das ferramentas empregadas. Os resultados mostraram que a broca especial resultou em acabamento e desvio dimensionais tão bons quanto aqueles produzidos pelo alargador. IT6 foi á qualidade obtida pelo alargador após furação com sobremetal de 0,5 mm, enquanto que a broca especial produziu qualidade IT8. O desgaste da broca especial foi menor que aquele dos alargadores, sendo que para estes últimos ocorreu lascamento. Cerca de 29% de redução no tempo de usinagem foram obtidos com á broca especial, o que resultou em um custo de produção 33% menor. Em contrapartida os valores dos desvios de cilindricidade foram superiores aqueles obtidos com os alargadores nas condições investigadas. A microdureza não foi afetada pelos processos empregados. / Mestre em Engenharia Mecânica

No description available.

Sortases are cell-membrane associated cysteine transpeptidases that are essential for the assembly and covalent anchoring of certain surface proteins to the cell wall in Gram-positive bacteria. Thus, they play critical roles in virulence, infection and colonization by pathogens. Sortases have been classified as type A, B, C, D, E and F based on their phylogeny and the target-protein motifs that they recognize. Sortase A (SrtA) enzymes participate in cell wall anchoring of proteins involved in bacterial adhesion, immune evasion, internalization, and phage recognition and in some cases pili formation. SrtA substrates are characterised by the presence of a C-terminal cell wall sorting signal as LPXTG motif, followed by a stretch of hydrophobic residues and a positively charged tail. Experimental and bioinformatics studies show that class A sortases are housekeeping as well as virulence determining proteins. Hence, Sortase A enzymes are considered as promising antibacterial drug targets, particularly because many organisms are developing multi-drug resistance behaviour. SrtA adopts an eight-stranded β-barrel structure and the overall fold is conserved among the sortase isoforms, with some modifications. The thesis candidate has determined the three dimensional (3D) crystal structures of wild-type and active site mutant of Sortase A from Streptococcus pneumoniae R6 strain by using X-ray diffraction method. The wild-type enzyme crystallized in P21 space group whereas active site cysteine mutant crystallized in C2 space group. In both the cases, N-terminal 81 residue deletion constructs (ΔN81) were used for crystallization. Uncommonly, both the structures showed a phenomenon of domain-swapping which resulted in the protein adopting a domain-swapped dimeric form. Two such dimers in wild-type protein and three dimers in mutant protein were observed in the asymmetric unit. To the best of our knowledge, our work reveals for the first time the occurrence of domain-swapping in sortase superfamily. Experimental techniques like size-exclusion chromatography, native-PAGE, analytical centrifugation and thiol cross-linking (carried out in our collaborator’s laboratory at National Institute of Immunology (NII), New Delhi, India) of functionally active wild-type SrtA from S. pneumoniae showed dimerization as well as domain-swapping in solution state. These results support the possibility that the protein indeed exists in a domain-swapped dimeric form and the determined structure is not the result of crystal packing artifact but is physiologically relevant as well. The work done by the thesis candidate covering crystallization of both, the active and inactive protein constructs, their structure determination using molecular replacement method, detailed structural analyses, structural comparisons with known SrtA structures and new structural findings are described in from Chapter 2 to Chapter 4. Based on the SrtA crystal structure the author of the thesis has also proposed various point mutations which are likely to disrupt domain– swapping and result in loss of dimer formation. In addition, as a part of the ongoing project in our laboratory, molecular dynamics studies of these domain-swapped dimers containing two sets of active site residues facing each other in a very compact volume have been initiated to understand substrate binding, which in future could lead to inhibitor design. Apart from the crystal structure analyses of SrtA structures, the author of the thesis has also carried out systematic crystal structure investigation of dipeptides and pentapeptides containing non-standard amino acids (ΔPhe, Aib and β-amino acids) along with computational studies. Conformationally restricted α,β-dehydrophenylalanine residue (ΔF) and α-aminoisobutyric acid (Aib) have been incorporated in highly amyloidogenic human Islet Amyloid Polypeptide (hIAPP) fragments. Amyloid deposits, observed in a vast majority of Type 2 diabetic patients, are primarily on account of misfolding and aggregation into fibrils of hIAPP, a 37 residue endocrine hormone secreted by pancreatic β-cells. It has been suggested that intermediates produced in the process of fibrillization are toxic to insulin producing β-cells. Hence, the inhibition of misfolding of hIAPP that involves structural transition from its native state (coil and/or helical and/or transient helical conformation) to β-sheet conformation, could be a possible strategy to mitigate Type 2 Diabetes Mellitus (T2DM). All the peptides discussed in this thesis were synthesized in our collaborator, Prof. V. S. Chauhan’s laboratory at the International Centre for Genetic Engineering and Biotechnology (ICGEB), New Delhi, India. In this work, author of the thesis has designed short peptides containing helicogenic residue, α,β-dehydrophenylalanine (ΔF) and determined their 3D crystal structures. It was found that pentapeptides, FGA∆FL and FGA∆FI act as inhibitors of hIAPP fibrillization. As revealed by crystal structure analysis, both the peptides have similar backbone conformation consisting of a ‘nest’ motif, which is an anion receptor. Molecular docking suggested that both the pentapeptides interact with the hIAPP20-27 segment, stabilizing the hIAPP in helical form by shielding the core aggregation initiation region. This reduces the possibility of oligomerization, formation of toxic intermediates and subsequently the transition to β-structure and fibrillization. Thus, the crystal structures of pentapeptide inhibitors together with computational docking studies suggest an atomic level model of the possible mode of action by which the FGAΔF(L/I) peptides manifest their fibrillization inhibition activity and this could be of value in the design of a new class of amyloid inhibitors. In another peptide design, L→U (Aib) mutation was done in core fibrillization region ANFLV i.e. hIAPP13-17. The resulting mutant peptide ANFUV as well as native fragment ANFLV was crystallized and their 3D crystal structures were determined. ANFLV crystallized in two space groups C2 and P2 adopting extended conformation. Crystal packing of ANFLV in both the crystals shows parallel beta sheet arrangement which is favoured and strengthened by hydrogen bonding between asparagine side chains of Asn-Asn pair each located in neighbouring parallel beta-strands. Hydrogen bonded Asn-Asn residue pairing in parallel beta-strands suggests its significant contribution during hIAPP fibril formation. The substitution L→U abolished its fibrillization property and the structure of ANFUV was solved by direct methods in P21 space group. The occurrence of β-bulge in ANFUV induced by Aib, as observed in crystal packing, suggests that Aib acts as a β-breaker through β-bulge inducing property in the highly amyloidogenic hIAPP segment. β-bulge forming property, an attribute of Aib as β-breaker may be responsible for the curtailment of fibrillization potential of the peptide in which the residue was incorporated. The aim of the anti-amyloid work is to design potent anti-fibrillization peptides and the work is important to design peptide based drugs to fight type II diabetes. The utilization of ΔPhe in the molecular self-assembly offers an added benefit in terms of variety and stability. Taking advantage of the conformation constraining property of ΔPhe residue, its incorporation in dipeptide molecules has been probed. The author has studied nanotube formation through molecular self-assembly, involving two classes of non¬standard amino acids i.e. ΔF and β-amino acids. FΔF in D-form, L-form and DL-mixture crystallized in different space groups forming rectangular/hexagonal channels constituting different channel dimensions. Recently, the application of FΔF nanotubes have been demonstrated in controlled drug delivery, showing the relevance of the work in health care. Another class of dipeptides containing β-amino acids (β-FF, β-FΔF, β-AΔF, β-VΔF, β¬LΔF, β-IΔF, and β-LF) was also explored for the self-assembled nanotube formation. These β-peptides were crystallized and their 3D structures were determined solely by the author of the thesis. Except the β-AΔF & β-LΔF, these peptides self-assemble and form rectangular/ hexagonal channels. Structures of ΔF and β-amino acid containing dipeptides forming ordered nanotubes through self-assembly are detailed in Chapters 8 and 9 in the thesis. Overall, the author of the thesis has crystallized and determined structures of more than twenty peptides. Experimentally, β-peptide nanotubes were observed to encapsulate drug molecules and thus might be useful as a drug delivery system. In the present thesis crystal structures of the following designed peptide sequences (including one natural sequence ANFLV) are reported in detail. Table 1 Peptide sequence Representation Length Discussed in 1. Phe-Gly-Ala-ΔPhe-Leu FGAΔFL 5 Chapter 6 2. Phe-Gly-Ala-ΔPhe-Ile FGAΔFI 5 Chapter 6 3. Ala-Asn-Phe-Leu-Val (2 forms) ANFLV_P2, ANFLV_C2 5 Chapter 7 4. Ala-Asn-Phe-Aib-Val ANFUV 5 Chapter 7 5. LPhe-ΔPhe (2 forms) LFΔF1 , LFΔF2 2 Chapter 8 6. DPhe-ΔPhe DFΔF 2 Chapter 8 7. DLPhe-ΔPhe DLFΔF 2 Chapter 8 8. LTyr-ΔPhe LYΔF 2 Chapter 8 9. LSer-ΔPhe LSΔF 2 Chapter 8 10. Boc-D,LPhe-ΔPhe Boc-DLFΔF 2 Chapter 8 11. Cbz-D,LPhe-ΔPhe Z-DLFΔF 2 Chapter 8 12. D,LMet-ΔPhe DLMΔF 2 Chapter 8 13. β-Phe-ΔPhe β-FΔF 2 Chapter 9 14. β-Phe-Phe β-FF 2 Chapter 9 15. β-Val-ΔPhe β-VΔF 2 Chapter 9 16. β-Ile-ΔPhe β-IΔF 2 Chapter 9 17. β-Leu-ΔPhe β-LΔF 2 Chapter 9 18. β-Leu-Phe β-LF 2 Chapter 9 19. β-Ala-ΔPhe β-AΔF 2 Chapter 9 20. Cyclo(Phe-ΔPhe) DKP-FΔF 2 Appendix C 21. Cyclo(Ile-ΔPhe) DKP-IΔF 2 Appendix C 22. Cyclo(Cha-Cha) DKP-ChaCha 2 Appendix C 23. Cyclo(Cha-Phe) DKP-ChaF 2 Appendix C 24. Cyclo(Cha-ΔPhe) DKP-ChaΔF 2 Appendix C 25. Cyclo(S-tritylCys-ΔPhe) DKP-CΔF 2 Appendix C Most of the dipeptides, except the N-terminal protected dipeptides, cyclic dipeptides (i.e. DKPs) and LSΔF, were found in the zwitterionic conformation and out of these, ten dipeptides resulted in tubular structures of dimensions in the nanoscale range. The thesis is organized into nine chapters and five appendices. Chapter 1 is an introduction to the work presented in the thesis, while Chapter 2, Chapter 3 and Chapter 4 describe the crystallographic work on the protein Sortase A. Chapter 5 is an introduction to the non-standard amino acids used for peptide designs and Chapter 6, Chapter 7, Chapter 8, Chapter 9 and Appendix C describe the crystallographic work on peptides. Chapter 1 starts with a general introduction to the Gram-positive bacteria containing sortase enzymes, and the bacterial cell-wall where sortase catalyzed proteins get attached for implicating their virulence during host-pathogen interactions. Pneumococcal diseases mostly affect children and their count has been observed to be higher than the combined total cases of malaria, AIDS and tuberculosis in child population worldwide. The chapter describes different virulence factors of S. pneumoniae out of which many are proteins. Among these, LPXTG containing proteins, which are the prime substrates of the sortase enzymes, are discussed in detail. Sortase enzymes, their classification and their structural studies with conserved ‘Sortase fold’ are discussed elaborately. A brief mention is made about the enzymatic activity of Sortase A to understand the transpeptidation mechanism. To appreciate the biomedical and biotechnological importance of the sortase enzyme, some potential applications of Sortase A are detailed in this chapter. A section is dedicated to describe the protein in the present study 'Sortase A from Streptococcus pneumoniae'. At the end, the scope of the present work, comprising of both protein and peptide crystallography, is presented. Chapter 2 begins with a brief account of the sequence analysis of Sortase A from S. pneumoniae and phylogenetic analysis of the sortase superfamily enzymes, followed by the details of protein purification & crystallization of two different constructs, wild-type SrtA from S. pneumoniae (Spn-∆N59SrtAWT and Spn-∆N81SrtAWT) as well as that of an active site cysteine mutant (Spn-∆N81SrtAC207A). This chapter includes X-ray intensity data collection of both types of crystals and data processing. Sortases are membrane anchored enzymes and therefore their expression as a full-length protein is a difficult task. Hence, the deletion of N-terminal transmembrane region from the enzyme is crucial for expression in its soluble form and is important for its successful crystallization. Thus, two wild-type constructs of S. pneumoniae sortase A, ∆N59SrtAWT (N-terminal 59 residue deletion) and ∆N81SrtAWT (N-terminal 81 residue deletion), and one active site mutant ∆N81SrtAC207A (N-terminal 81 residue deletion & active site Cys207 to Ala mutation) were cloned, expressed and purified. Cloning, expression and purification of the protein were done at the laboratory of our collaborator Prof. Rajendra P. Roy, Cell biology lab-II, National Institute of Immunology (NII), New Delhi, India. Crystallization of Spn-∆N59SrtAWT (~23 kDa) construct was initiated by manual screening using sparse matrix conditions from Hampton research. Initial trials were set up by following hanging-drop vapour diffusion method. Spn-∆N59SrtAWT construct crystallized in diamond, needle, rod and wedge-shaped crystal forms in more than one crystallization condition but they failed to diffract. Further trials were set up in microbatch plates that resulted in diamond-shaped crystals again, which diffracted up to a maximum of 4.0 Å resolution. Sequence comparison of the present construct was performed to modify the construct to achieve better diffraction. Thus, we made modifications in the Spn¬∆N59SrtAWT construct by deleting additional 22 residues at the N-terminal (i.e. total 81 residues deletion in the original sequence from the N-terminal) similar to SrtA from S. pyogenes. Hence, Spn-∆N81SrtAWT construct was prepared. For further crystallization experiments, we used the new construct Spn-∆N81SrtAWT. Similar to Spn-∆N59SrtAWT construct, crystallization set up for Spn-∆N81SrtAWT were done in microbatch plates at 293 K by using the Hampton conditions. During the crystallization set up, protein concentration was varied from 6-30 mg/ml. Notably, the protein crystals grown with 25 mg/ml protein concentration diffracted very well. Thus increasing the protein concentration helped to improve diffraction quality. Crystals obtained in Index-88 condition (0.2 M tri-ammonium citrate and 20% (w/v) PEG 3350, pH 7.0) diffracted up to 2.9 Å. Additive screen was used to improve its diffraction quality. This time many diffracting crystals were obtained and the best rod-shaped crystals grown in additive screen-79 (40% v/v (±)-1,3-butanediol) diffracted well up to 2.70 Å at home source. Thus, Spn-ΔN81SrtAWT crystallized at protein concentration of 25 mg ml-1 (in 10 mM Tris buffer, pH 7.5; 2 mM β-mercaptoethanol) with a condition containing 0.2 M tri-ammonium citrate and 20% (w/v) PEG 3350, pH 7.0, along with 40% v/v (±)¬1,3-butanediol as an additive agent by using microbatch-under-oil crystallization method. The chapter also includes crystallization of active site mutant Cys207Ala of ∆N81SrtAWT from S. pneumoniae (Spn-∆N81SrtAC207A). Spn-∆N81SrtAC207A mutant crystallized as a beautiful rectangular block type crystal (with a diffraction up to 2.7 Å at home source and up to 2.48 Å at synchrotron) at protein concentration of 25 mg ml-1 (in 10 mM Tris buffer, pH 7.5; 2 mM β-mercaptoethanol) with a condition containing 0.2 M tri-ammonium citrate and 20% (w/v) PEG 3350, pH 7.0, along with 1.0 M guanidine hydrochloride as an additive agent by using microbatch-under-oil crystallization method. Data collection was done on home-source diffraction facility for both the crystals however; mutant data in better resolution was collected by the author of the thesis at BM-14 beamline at ESRF, Grenoble, France. Thus, two crystals of SrtA, wild-type (Spn-∆N81SrtAWT) and its C207A mutant (Spn-∆N81SrtAC207A) were indexed satisfactorily in two space groups and their cell parameters are given in the following table 2. Table 2 Protein Space group a (Å) b (Å) c (Å) β (°) X-ray source Spn-∆N81SrtAWT P21 66.94 103.45 74.87 115.65 Home source Spn-∆N81SrtAC207A C2 155.57 113.33 81.34 90.80 Synchrotron The quality of both the data sets was assessed by SFCHECK and none of them showed twinning. Thus, the data sets collected were found appropriate and useful for structure determination as discussed in Chapter 3. Chapter 3 details the structure determination of Sortase A from S. pneumoniae for a wild-type construct (Spn-ΔN81SrtAWT) and for an active site cysteine mutant construct (Spn-ΔN81SrtAC207A). Sortase A from S. pyogenes was used as a search model in the molecular replacement (MR) method and a single solution for each data set was obtained through PHASER program. It resulted in four-molecules in wild-type sortase structure and six-molecules in the mutant structure in the respective crystal asymmetric unit. Iterative model building and structure refinement revealed a clear case of domain-swapping as observed in the electron density map. Finally, in the asymmetric unit of wild-type structure and in mutant protein structure two and three domain-swapped dimers were located, respectively. Simulated annealing and TLS refinement resulted in the protein structure with best refinement statistics. All these are elaborately discussed in Chapter 3. The last round of refinement of Spn-ΔN81SrtAWT converged to Rwork = 18.10% and Rfree = 23.39 % for 25152 unique reflections in the resolution range 30.7-2.7 Å whereas for Spn¬ΔN81SrtAC207A structure these parameters converged to Rwork = 18.25% and Rfree = 22.39% for 50010 unique reflections in the resolution range 47.15-2.48 Å. Chapter 4 describes the wild-type (Spn-ΔN81SrtAWT) as well as mutant (Spn¬ΔN81SrtAC207A) structures of Sortase A. The structure of Sortase A is not found in its commonly observed monomeric form but occur in a domain-swapped dimeric form. There are two dimers in Spn-ΔN81SrtAWT and three in Spn-ΔN81SrtAC207A as observed in the asymmetric unit. Each dimer contains two characteristic 8-stranded beta-barrel folds i.e. ‘sortase fold’ which is unique to the sortase superfamily. Unlike the structure of SrtA from other organisms known so far, the monomer does not form the 8-stranded beta-barrel all by itself. One monomer exchanges the β7 and β8 strands with the other monomer having β1 to β6 strands, thereby forming a complete 8-stranded β-barrel fold and such kind of two complete folds are present in each dimer. Because of the mutual swapping of strands between two monomers in a dimer, the dimer thus formed is defined as a domain-swapped dimer. This is the first time we have observed Sortase A structure in the domain-swapped dimeric form and is also the first example of domain-swapping in the sortase superfamily. Interestingly, all the catalytic residues (His141, Cys207 and Arg215) in each sortase fold in the swapped dimer lie at the secondary interface (open interface) generated by domain-swapping. Catalytic R215 (in one fold) interacts with D209 residue (in other fold of same dimer) through salt bridge interactions. Each dimer contains two pairs of such residues at the secondary interface but only one pair shows this kind of interaction. R215 (B-chain) interacts with D209 (A-chain) in AB dimer whereas R215 (D-chain) interacts with D209 (C-chain) in CD dimer. Asymmetry in the catalytic residues for their orientations and observed interactions at the secondary interface was evidenced. These active site residues were seen buried to a great extent except Arg215 which is slightly better exposed. It was difficult to find the exact substrate-binding pocket to approach the catalytic Cys207. However, biochemical and biophysical analyses (done at NII, New Delhi) provided strong evidence for the existence of the swapped-dimeric form at physiological pH as well. The enzyme exists with an equilibrium between its monomeric and dimeric forms, and the dimeric population is the most active species of the functionally active enzyme. An important role of Glu208 (in all the chains of two dimers; e.g. Chain A) was seen in the catalytic site where its side chain wobbles between His141 and H142 (both in Chain B) residues for interaction. Due to such kind of interactions the backbone conformation between C207-E208 (Chain B) shows variability, and coordinates the distance between His141 (ND1, Chain A) and Cys207 (SG, Chain B) each belonging to opposite chains in a swapped-dimer. The nature of side chain conformations of Glu208 in all the four sets of active site residues (in wild-type as well as in cysteine mutant structure) indicates that its movement presumably regulates thiolate-imidazolium acid-base pair formation which is a crucial condition for the sortase function where cysteine thiolate acts as nucleophile. Based on the crystal structure, the thesis candidate has suggested several mutants which might disrupt domain-swapping pointing to future studies on the system. Domain movement analyses by using HingeProt and DynDom servers indicate that the two-sortase folds joined with hinge loops in each dimer may show twist movement around the hinge axis. Possibly, such motion will affect the secondary interface covering active site residues and may allow increasing the exposure of the catalytic residues to perform catalysis. Presumably, such kind of domain movements may play a key role for the unique kind of regulatory mechanism for transpeptidase activity in sortase enzymes. However, more study has to be done to explore the role of these possibilities, if any, in the enzyme function and its regulation. Chapter 5 provides an introduction to non-standard amino acids, their sources and their uses in de novo peptide design; this is followed by a description of outcomes of structural investigations of modified peptides and their applications in various fields of medical and material science. Specifically, α, β-dehydrophenylalanine (ΔPhe), α-aminoisobutyric acid (Aib) and β-amino acids are discussed and their structures and conformational preferences are highlighted for their use in naturally occurring peptides or peptide fragments. Chapter 6 begins with an introduction to the human Islet Amyloid Polypeptide (hIAPP), which is an amyloidogenic protein and considered to be an important protein constituent of the amyloid plaques in pancreatic beta-cells in Type 2 diabetes patients. Therefore, fibrillization inhibition of hIAPP is considered as an important therapeutic approach to combat Type 2 Diabetes Mellitus (T2DM). In this chapter, the author of the thesis describes an approach to design peptide based inhibitors of hIAPP fibrillization using non¬standard amino acid ΔPhe (α,β-dehydrophenylalanine) residue. The first designed inhibitor has the sequence origin from hIAPP23-27 and it was developed by replacing I→ΔF (i.e. β¬favouring residue to helical conformation favouring) which resulted in FGAΔFL peptide. Fibrillization inhibition studies were done by co-incubation of hIAPP and FGAΔFL in 1:5 molar ratio and monitored by electron microscopy and thioflavin T binding assay that showed ~75% fibrillization inhibition. It suggested that the inhibitor is working effectively and thus the author determined its crystal structure by X-ray diffraction method. Peptide synthesis and experimental studies like electron microscopy and Thioflavin T binding assay were done in our collaborator’s laboratory at ICGEB, New Delhi, India. Subsequently a sequence similar peptide FGAΔFI was also designed by mutating L→I in the first inhibitor sequence. The resulting peptide FGAΔFI showed ~70% fibrillization inhibition. Following this success, crystal structures of both peptides were determined. FGAΔFL crystallized in P212121 space group whereas FGAΔFI crystallized in P21 space group. Though it was not anticipated, crystal structure analysis revealed that FGAΔFL and its analogue FGAΔFI harbour the anion receptor ‘nest’ motif. Both peptides dock with the helical form of hIAPP which may contribute to the inhibitory function of the peptides through their interaction with hIAPP in the core fibrillization region. These peptides effectively inhibit hIAPP fibrillization in vitro and it seems that these are unique examples of ‘nest-motif’ containing peptides that inhibit fibrillization. We also propose a model for fibrillization inhibition by these peptides; this has been published in Chemical Communications, a journal published by the Royal Society of Chemistry (RSC) and its reprint is enclosed within the thesis. In general, the approach described in the chapter may be applicable to target helices or helical intermediates and could be utilized in developing inhibitors useful, apart from T2DM, in other amyloid diseases including Alzheimer’s disease and Parkinson’s disease. Table 3 Peptide Crystal system and space group Unit cell details X-ray data Structure solution and refinement Agreement factor FGAΔFL Orthorhombic, P212121 a=8.9951 (9) Åb=13.0144 (12) Åc=27.7521 (24) ÅV=3248.82 (5) Å3 Z=4 Mo Kα(λ=0.71073Å) 4703 Unique reflections 2581 [|Fo| > 4σ (|Fo|)] Direct methods: SHELXS97 & SHELXL97 5.95 % for [|Fo| > 4σ (|Fo|)] FGAΔFI Monoclinic, P21 a=8.9951 (9) Åb=13.0144 (12) Åc=27.7521 (24) Å β=92.637 (2)°V=935.59 (2) Å3 Z=2 Mo Kα(λ=0.71073Å) 4024 Unique reflections 2612 [|Fo| > 4σ (|Fo|)] Direct methods: SHELXS97 & SHELXL97 5.02 % for [|Fo| > 4σ (|Fo|)] Chapter 7 describes another important but less studied core fibrillization fragment of hIAPP (hIAPP13-17) different than the hIAPP23-27 discussed in the previous chapter. It also discusses the development of fibrillization inhibitor design from this segment. The fragment hIAPP13-17 i.e. ANFLV crystallized in two space groups; C2 with one molecule in the asymmetric unit and P2 with two molecules in the asymmetric unit. In these structures, ANFLV peptide shows fully extended conformation i.e. a β-conformation. Crystal packing shows parallel β-sheet arrangement with the involvement of dry ‘steric-zippers’. The peptide prefers cross-strand Asn-Asn residue pair by side chain hydrogen bonding and is discussed in comparison with a few crystal structures of hIAPP fragments, solved by Eisenberg’s group, containing Asn residue in their sequence. It is observed that if the Asn is located in the sequence between two terminal residues the peptide will arrange itself in parallel beta sheet. This supports a structural model of hIAPP fibril in parallel beta sheet arrangement as the hIAPP sequence contains several Asn residues. To develop an inhibitor from ANFLV, a partial success was achieved where the Leu → Aib mutant i.e. ANFUV was developed. ThT (Thioflavin T) and TEM (Transmission electron microscopy) results show that the mutant peptide does not fibrilize on its own. This strongly supports the fact that the native peptide (ANFLV) lost its inherent fibrillization characteristic with the introduction of Aib in place of Leu i.e. the resultant mutant ANFUV is a non-fibrillizing peptide. The logic behind the development was to retain ANF in the same extended conformation and then break the β-strand with β-breaker residues. The structure of ANFLV showed parallel beta-sheets along with the additional side chain-side chain hydrogen bonding in the same direction as the fibril axis. Thus, we retained the ANF region to keep the sticky segment in the design and then Leu was mutated to Aib, a known β-breaker, to alter backbone conformation. The crystal structure of the peptide ANFUV resulted in the similar ANF region in beta conformation and Aib in helical conformation. Interestingly, in this situation the conformation of Aib develops a beta-bulge observed in the crystal packing and this bulge structure probably turned the peptide to have non-fibrillizing characteristics. These results will be useful in designing peptide inhibitors by using U as a beta breaker to inhibit hIAPP fibrillization. Table 4 Peptide Crystal system and space group Unit cell details X-ray data Structure solution and refinement Agreement factor ANFLV1 Monoclinic, C2 a=36.1350 (20) Åb=4.8050 (10) Åc=19.4190 (20) Å β=98.644 (5)°V=3333.40 (27) Å3 Z=4 Synchrotron (λ=0.77490 Å) 1982 Unique reflections 1825 [|Fo| > 4σ (|Fo|)] Direct methods: Sir92 & SHELXL97 11.71% for [|Fo| > 4σ (|Fo|)] ANFLV2 Monoclinic, P2 a=18.7940 (80) Åb=4.7970 (10) Åc=35.4160 (50) Å β=103.929 (10)°V=3099.03 (81) Å3 Z=4 Synchrotron (λ=0.77490 Å) 2651 Unique reflections 2580 [|Fo| > 4σ (|Fo|)] Direct methods: Sir92 & SHELXL97 15.39% for [|Fo| > 4σ (|Fo|)] ANFUV Monoclinic, P21 a=10.8140 (22) Åb=9.1330 (18) Åc=16.7540 (34) Å β=107.960 (30)°V=1574.07 (161) Å3 Z=2 Synchrotron (λ=0.97918 Å) 1426 Unique reflections 1398 [|Fo| > 4σ (|Fo|)] Direct methods: SHELXS97 & SHELXL97 5.45% for [|Fo| > 4σ (|Fo|)] Chapter 8 elaborates the self-assembly of α-dipeptides containing conformationally constrained achiral amino acid, α,β-dehydrophenylalanine (ΔF). The structural polymorphism in LFΔF peptide and the resulting self-assembly are discussed. Its D-isomer (DF∆F) and its racemic mixture (DLF∆F) are also discussed as these peptides self-assemble to give channel-forming assemblies. In addition to LFΔF, crystal structures of LYΔF, DLMΔF and LSΔF peptides and their self-assemblies are presented as well. Except DLMΔF xi and N-terminal protected DLFΔF (Boc-DLF∆F and Z-DLF∆F) peptides, the other dipeptides discussed in this chapter resulted in tubular structures of nanoscale dimensions through molecular self-aggregation. Table 5 Peptide Crystal system and space group Unit cell details X-ray data Structure solution and refinement Agreement factor LFΔF1 Hexagonal, P65 a=23.1873(24) Åb=23.1873(24) Åc=5.5260(8) ÅV=2573.01(5) Å3 Z=6 Mo Kα(λ=0.71073Å) 3489 Unique reflections 2915 [|Fo| > 4σ (|Fo|)] Direct methods: SHELXS97 & SHELXL97 6.19% for [|Fo| > 4σ (|Fo|)] LFΔF2 Monoclinic, P21 a=5.5739(2) Åb=13.1383(4) Åc=13.5816(4) Å β=96.137(2)°V=988.90(2) Å3 Z=2 Mo Kα(λ=0.71073Å) 4865 Unique reflections 3402 [|Fo| > 4σ (|Fo|)] Direct methods: SHELXS97 & SHELXL97 4.35% for [|Fo| > 4σ (|Fo|)] DFΔF Orthorhombic, P21212 a=13.1690(21) Åb=25.3673(40) Åc=5.5622(9) ÅV=1858.12(5) Å3 Z=4 Mo Kα(λ=0.71073Å) 4370 Unique reflections 3426 [|Fo| > 4σ (|Fo|)] Direct methods: SHELXS97 & SHELXL97 4.44% for [|Fo| > 4σ (|Fo|)] DLFΔF Monoclinic, P21/c a=5.5392(14) Åb=26.0376(55) Åc=13.1839(27) Å β=90.278(16)°V=1901.46(8) Å3 Z=4 Mo Kα(λ=0.71073Å) 2051 Unique reflections 1264 [|Fo| > 4σ (|Fo|)] Direct methods: SHELXS97 & SHELXL97 7.08% for [|Fo| > 4σ (|Fo|)] LYΔF Hexagonal, P65 a=23.5523(4) Åb=23.5523(4) Åc=5.5183(1) ÅV=2650.96(1) Å3 Z=6 Mo Kα(λ=0.71073Å) 2746 Unique reflections 1871 [|Fo| > 4σ (|Fo|)] Direct methods: SHELXS97 & SHELXL97 3.91% for [|Fo| > 4σ (|Fo|)] LSΔF Monoclinic, P21 a=5.2998(20) Åb=9.6732(30) Åc=14.1827(57) Å β=95.604(27)°V=723.62(20) Å3 Z=2 Mo Kα(λ=0.71073Å) 1978 Unique reflections 1558 [|Fo| > 4σ (|Fo|)] Direct methods: SHELXS97 & SHELXL97 13.59% for [|Fo| > 4σ (|Fo|)] DLMΔF Monoclinic, P21/c a=9.9032(5) Åb=8.6675(4) Åc=34.0283(18) Å β=90.088(3)°V=29

abstract: Universities and colleges in the United States (U.S.) are in a period of rapid transformation. Driven by the need for an educated workforce, higher education institutions are responding to rapid innovation, globalization, economic realities, and sociodemographic shifts. Simultaneously, extensive educational online networks connect millions of people worldwide enable learning and knowledge sharing beyond what society has experienced to date. In light of technological advancements, the preservation and presentation of certain ideals that undergird academia and the communication and application of knowledge are undergoing dramatic change. Within higher education, this is both a challenge and an opportunity to re-envision the commitment to educate the public. This research discusses potential forms of this redesign and how it can build upon and depart from previous iterations of higher education. How colleges and universities will adapt to become more relevant, engaging, and accessible is a pressing question that must be addressed. Using case studies focused on creating sustainability education materials, this dissertation develops knowledge related to three interconnected areas of study that will contribute to redesigning higher education through participatory action research methodology. First, higher education has a civic responsibility to provide new ways of thinking, being, and doing globally and providing more access to education to broader society, especially through public research institutions. Second, with a vast array of available learning materials, higher education should invest in elegantly-designed experiences consisting of well-reasoned, meticulously-curated, and high-quality content that is aesthetically appealing, engaging, and accessible to a broad audience. Third, as universities transition from the gatekeepers of knowledge to the connectors of knowledge, they also need to ensure that a coherent mission is articulated and invested in by stakeholders to create an intentionally beneficial transformational effort. The transformation of higher education toward a more inclusive learning environment through new ways of thinking and elegantly-designed learning experiences will serve to improve our learning institutions. As part of the necessary core for an educated democracy, higher education institutions must strive to create a more equitable, inclusive, and diverse society. / Dissertation/Thesis / Doctoral Dissertation Human and Social Dimensions of Science and Technology 2020

The main aim of the thesis was the liquefaction of Únanovka streamflow under the Těšetice dam with the focus on suitable flood protection of the villages of Těšetice, Bantice and Práče.The thesis evaluates the existing condition of the Únanovka streamflow in the river mileage 1.6593 – 6.4103. Calculation of the streamflow capacity was carried out per 1D mathematical model in the HEC-RAS programme for the selected N-year flow from the ČHMÚ hydrological study. Final value of the improved drainage from the dam was determined together with the possible value of water collection. Within the thesis the hydro-ecological monitoring of the watercourse was carried out (HEM analysis 2014).

Sweden's population is constantly growing and so are the larger cities. Meanwhile thousands of buildings in the countryside are unused and forgotten. Recent world events have resulted in more opportunities for remote work, enabling more freedom in choosing where to live.The aim of this study is to exemplify how the change of an abandoned building can contribute to social sustainability and development of rural areas. The suggestions presented in the report should be applicable in accordance with current regulations, such as Plan- och bygglagen. Therefore a presentation of relevant laws is included in the study.The exemplification in this study is presented in the form of a design for a repurposed school-building in a small society near Älmhult. The design allows the space to be used for public activities and greenery. This is in accordance with theories and answers from interviews, which were conducted with a selection of people relevant to the subject, regarding how to make rural areas more attractive.

New Genre Public Art was originally defined by Suzanne Lacy in 1991 as an activist approach to the public; it was a type of public art that was often created outside the institutional structure which brought the artist into direct engagement with the audience, while addressing social and political issues. In 1993, the public art exhibition ’Culture in Action’, curated by Mary Jane Jacob, marked a conceptual shift from static to dynamic public art. The exhibition is considered a landmark event in the development of public art as it was among the first projects to frame communities as the structure and content of its art.During the past decade (2010–2020), urban development has become incorporated as an integral part of the work of the Public Art Agency Sweden, and the agency have established their own curatorial department in order to curate and produce their own public art exhibitions. As Public Art Agency Sweden is a State agency, their work is largely determined by official policies formulated by the Swedish government. This thesis analyzes the contemporary policies of urban public art by conducting an interdisciplinary critical discourse analysis that merges art history, curatorial– and urban studies, in order to trace the influence of discourse to how Public Art Agency Sweden has operated within this intersection during the last decade––ultimately to discuss what the Swedish policies on public art strive to achieve and the risks, ethics and responsibilities of the emerging field of urban, context-based curatorship.