Matching Items (188)
Description
Membrane proteins are very important for all living cells, being involved in respiration, photosynthesis, cellular uptake and signal transduction, amongst other vital functions. However, less than 300 unique membrane protein structures have been determined to date, often due to difficulties associated with the growth of sufficiently large and well-ordered crystals. This work has been focused on showing the first proof of concept for using membrane protein nanocrystals and microcrystals for high-resolution structure determination. Upon determining that crystals of the membrane protein Photosystem I, which is the largest and most complex membrane protein crystallized to date, exist with only a hundred unit cells with sizes of less than 200 nm on an edge, work was done to develop a technique that could exploit the growth of the Photosystem I nanocrystals and microcrystals. Femtosecond X-ray protein nanocrystallography was developed for use at the first high-energy X-ray free electron laser, the LCLS at SLAC National Accelerator Laboratory, in which a liquid jet would bring fully hydrated Photosystem I nanocrystals into the interaction region of the pulsed X-ray source. Diffraction patterns were recorded from millions of individual PSI nanocrystals and data from thousands of different, randomly oriented crystallites were integrated using Monte Carlo integration of the peak intensities. The short pulses ( 70 fs) provided by the LCLS allowed the possibility to collect the diffraction data before the onset of radiation damage, exploiting the diffract-before-destroy principle. At the initial experiments at the AMO beamline using 6.9- Å wavelength, Bragg peaks were recorded to 8.5- Å resolution, and an electron-density map was determined that did not show any effects of X-ray-induced radiation damage. Recently, femtosecond X-ray protein nanocrystallography experiments were done at the CXI beamline of the LCLS using 1.3- Å wavelength, and Bragg reflections were recorded to 3- Å resolution; the data are currently being processed. Many additional techniques still need to be developed to explore the femtosecond nanocrystallography technique for experimental phasing and time-resolved X-ray crystallography experiments. The first proof-of-principle results for the femtosecond nanocrystallography technique indicate the incredible potential of the technique to offer a new route to the structure determination of membrane proteins.
ContributorsHunter, Mark (Author) / Fromme, Petra (Thesis advisor) / Wolf, George (Committee member) / Levitus, Marcia (Committee member) / Arizona State University (Publisher)
Created2011
Description
With the increasing focus on developing environmentally benign electronic packages, lead-free solder alloys have received a great deal of attention. Mishandling of packages, during manufacture, assembly, or by the user may cause failure of solder joint. A fundamental understanding of the behavior of lead-free solders under mechanical shock conditions is lacking. Reliable experimental and numerical analysis of lead-free solder joints in the intermediate strain rate regime need to be investigated. This dissertation mainly focuses on exploring the mechanical shock behavior of lead-free tin-rich solder alloys via multiscale modeling and numerical simulations. First, the macroscopic stress/strain behaviors of three bulk lead-free tin-rich solders were tested over a range of strain rates from 0.001/s to 30/s. Finite element analysis was conducted to determine appropriate specimen geometry that could reach a homogeneous stress/strain field and a relatively high strain rate. A novel self-consistent true stress correction method is developed to compensate the inaccuracy caused by the triaxial stress state at the post-necking stage. Then the material property of micron-scale intermetallic was examined by micro-compression test. The accuracy of this measure is systematically validated by finite element analysis, and empirical adjustments are provided. Moreover, the interfacial property of the solder/intermetallic interface is investigated, and a continuum traction-separation law of this interface is developed from an atomistic-based cohesive element method. The macroscopic stress/strain relation and microstructural properties are combined together to form a multiscale material behavior via a stochastic approach for both solder and intermetallic. As a result, solder is modeled by porous plasticity with random voids, and intermetallic is characterized as brittle material with random vulnerable region. Thereafter, the porous plasticity fracture of the solders and the brittle fracture of the intermetallics are coupled together in one finite element model. Finally, this study yields a multiscale model to understand and predict the mechanical shock behavior of lead-free tin-rich solder joints. Different fracture patterns are observed for various strain rates and/or intermetallic thicknesses. The predictions have a good agreement with the theory and experiments.
ContributorsFei, Huiyang (Author) / Jiang, Hanqing (Thesis advisor) / Chawla, Nikhilesh (Thesis advisor) / Tasooji, Amaneh (Committee member) / Mobasher, Barzin (Committee member) / Rajan, Subramaniam D. (Committee member) / Arizona State University (Publisher)
Created2011
Description
The Kasturba Gandhi Balika Vidyalaya (KGBV) policy scheme launched in 2004 by the Ministry of Human Resource Development, the Government of India, aims to provide secondary level education (grade 6-8) for girls residing predominantly in minority communities, the Scheduled Caste (SC), the Scheduled Tribe (ST), and the Other Backward Caste (OBC). Since its launch, the Government of India established 2,578 KGBV schools in 27 states and union territories (UTs). The present study examines the new policy and its implementation at three KGBV schools located in rural villages of Uttar Pradesh (UP), India. The purpose was to analyze the Government of India's approach to increasing education opportunity and participation for educationally disadvantaged girls using the empowerment framework developed by Deepa Narayan. Observations at three schools, interviews with teachers and staff members of the implementation agency (i.e., Mahila Samakhya (MS)), and surveys administered to 139 teachers were conducted over a four month period in 2009. Adopting creative teaching approaches and learning activities, MS creates safe learning community which is appropriate for the rural girls. MS gives special attention to nurturing the girls' potential and empowering them inside and outside the school environment through social discussion, parental involvement, rigid discipline and structure, health and hygiene education, and physical and mental training. Interviews with the state program director and coordinators identified some conflicts within government policy schemes such as the Teacher-pupil ratios guidelines as a part of the programs for the universalization of elementary education. Major challenges include a high turnover rate of teachers, a lack of female teachers, a lack of provision after Class 8, and inadequate budget for medical treatment. Recommendations include promoting active involvement of male members in the process of girls' empowerment, making MS approaches of girls' education in rural settings standardized for wider dissemination, and developing flexible and strong partnership among local agencies and government organizations for effective service delivery.
ContributorsWatanabe, Miku (Author) / Fischman, Gustavo (Thesis advisor) / Wiley, Terrence (Committee member) / Mccarty, Teresa (Committee member) / Arizona State University (Publisher)
Created2011
Description
Molybdenum (Mo) is a key trace nutrient for biological assimilation of nitrogen, either as nitrogen gas (N2) or nitrate (NO3-). Although Mo is the most abundant metal in seawater (105 nM), its concentration is low (<5 nM) in most freshwaters today, and it was scarce in the ocean before 600 million years ago. The use of Mo for nitrogen assimilation can be understood in terms of the changing Mo availability through time; for instance, the higher Mo content of eukaryotic vs. prokaryotic nitrate reductase may have stalled proliferation of eukaryotes in low-Mo Proterozoic oceans. Field and laboratory experiments were performed to study Mo requirements for NO3- assimilation and N2 fixation, respectively. Molybdenum-nitrate addition experiments at Castle Lake, California revealed interannual and depth variability in plankton community response, perhaps resulting from differences in species composition and/or ammonium availability. Furthermore, lake sediments were elevated in Mo compared to soils and bedrock in the watershed. Box modeling suggested that the largest source of Mo to the lake was particulate matter from the watershed. Month-long laboratory experiments with heterocystous cyanobacteria (HC) showed that <1 nM Mo led to low N2 fixation rates, while 10 nM Mo was sufficient for optimal rates. At 1500 nM Mo, freshwater HC hyperaccumulated Mo intercellularly, whereas coastal HC did not. These differences in storage capacity were likely due to the presence in freshwater HC of the small molybdate-binding protein, Mop, and its absence in coastal and marine cyanobacterial species. Expression of the mop gene was regulated by Mo availability in the freshwater HC species Nostoc sp. PCC 7120. Under low Mo (<1 nM) conditions, mop gene expression was up-regulated compared to higher Mo (150 and 3000 nM) treatments, but the subunit composition of the Mop protein changed, suggesting that Mop does not bind Mo in the same manner at <1 nM Mo that it can at higher Mo concentrations. These findings support a role for Mop as a Mo storage protein in HC and suggest that freshwater HC control Mo cellular homeostasis at the post-translational level. Mop's widespread distribution in prokaryotes lends support to the theory that it may be an ancient protein inherited from low-Mo Precambrian oceans.
ContributorsGlass, Jennifer (Author) / Anbar, Ariel D (Thesis advisor) / Shock, Everett L (Committee member) / Jones, Anne K (Committee member) / Hartnett, Hilairy E (Committee member) / Elser, James J (Committee member) / Fromme, Petra (Committee member) / Arizona State University (Publisher)
Created2011
Biological and immunological characterization of plant-produced HIV-1 Gag/dgp41 virus-like particles
Description
Anti-retroviral drugs and AIDS prevention programs have helped to decrease the rate of new HIV-1 infections in some communities, however, a prophylactic vaccine is still needed to control the epidemic world-wide. Despite over two decades of research, a vaccine against HIV-1 remains elusive, although recent clinical trials have shown promising results. Recent successes have focused on highly conserved, mucosally-targeted antigens within HIV-1 such as the membrane proximal external region (MPER) of the envelope protein, gp41. MPER has been shown to play critical roles in the viral mucosal transmission, though this peptide is not immunogenic on its own. Gag is a structural protein configuring the enveloped virus particles, and has been suggested to constitute a target of the cellular immunity potentially controlling the viral load. It was hypothesized that HIV-1 enveloped virus-like particles (VLPs) consisting of Gag and a deconstructed form of gp41 comprising the MPER, transmembrane, and cytoplasmic domains (dgp41) could be expressed in plants. Plant-optimized HIV-1 genes were constructed and expressed in Nicotiana benthamiana by stable transformation, or transiently using a tobacco mosaic virus-based expression system or a combination of both. Results of biophysical, biochemical and electron microscopy characterization demonstrated that plant cells could support not only the formation of HIV-1 Gag VLPs, but also the accumulation of VLPs that incorporated dgp41. These particles were purified and utilized in mice immunization experiments. Prime-boost strategies combining systemic and mucosal priming with systemic boosting using two different vaccine candidates (VLPs and CTB-MPR - a fusion of MPER and the B-subunit of cholera toxin) were administered to BALB/c mice. Serum antibody responses against both the Gag and gp41 antigens could be elicited in mice systemically primed with VLPs and these responses could be recalled following systemic boosting with VLPs. In addition, mucosal priming with VLPs allowed for a robust boosting response against Gag and gp41 when boosted with either candidate. Functional assays of these antibodies are in progress to test the antibodies' effectiveness in neutralizing and preventing mucosal transmission of HIV-1. This immunogenicity of plant-based Gag/dgp41 VLPs represents an important milestone on the road towards a broadly-efficacious and inexpensive subunit vaccine against HIV-1.
ContributorsKessans, Sarah (Author) / Mor, Tsafrir S (Thesis advisor) / Matoba, Nobuyuki (Committee member) / Mason, Hugh (Committee member) / Hogue, Brenda (Committee member) / Fromme, Petra (Committee member) / Arizona State University (Publisher)
Created2011
Description
The cyanobacterium Synechocystis sp. PCC 6803 performs oxygenic photosynthesis. Light energy conversion in photosynthesis takes place in photosystem I (PSI) and photosystem II (PSII) that contain chlorophyll, which absorbs light energy that is utilized as a driving force for photosynthesis. However, excess light energy may lead to formation of reactive oxygen species that cause damage to photosynthetic complexes, which subsequently need repair or replacement. To gain insight in the degradation/biogenesis dynamics of the photosystems, the lifetimes of photosynthetic proteins and chlorophyll were determined by a combined stable-isotope (15N) and mass spectrometry method. The lifetimes of PSII and PSI proteins ranged from 1-33 and 30-75 hours, respectively. Interestingly, chlorophyll had longer lifetimes than the chlorophyll-binding proteins in these photosystems. Therefore, photosynthetic proteins turn over and are replaced independently from each other, and chlorophyll is recycled from the damaged chlorophyll-binding proteins. In Synechocystis, there are five small Cab-like proteins (SCPs: ScpA-E) that share chlorophyll a/b-binding motifs with LHC proteins in plants. SCPs appear to transiently bind chlorophyll and to regulate chlorophyll biosynthesis. In this study, the association of ScpB, ScpC, and ScpD with damaged and repaired PSII was demonstrated. Moreover, in a mutant lacking SCPs, most PSII protein lifetimes were unaffected but the lifetime of chlorophyll was decreased, and one of the nascent PSII complexes was missing. SCPs appear to bind PSII chlorophyll while PSII is repaired, and SCPs stabilize nascent PSII complexes. Furthermore, aminolevulinic acid biosynthesis, an early step of chlorophyll biosynthesis, was impaired in the absence of SCPs, so that the amount of chlorophyll in the cells was reduced. Finally, a deletion mutation was introduced into the sll1906 gene, encoding a member of the putative bacteriochlorophyll delivery (BCD) protein family. The Sll1906 sequence contains possible chlorophyll-binding sites, and its homolog in purple bacteria functions in proper assembly of light-harvesting complexes. However, the sll1906 deletion did not affect chlorophyll degradation/biosynthesis and photosystem assembly. Other (parallel) pathways may exist that may fully compensate for the lack of Sll1906. This study has highlighted the dynamics of photosynthetic complexes in their biogenesis and turnover and the coordination between synthesis of chlorophyll and photosynthetic proteins.
ContributorsYao, Cheng I Daniel (Author) / Vermaas, Wim (Thesis advisor) / Fromme, Petra (Committee member) / Roberson, Robert (Committee member) / Webber, Andrew (Committee member) / Arizona State University (Publisher)
Created2011
Description
ATP synthase is a large multimeric protein complex responsible for generating the energy molecule adenosine triphosphate (ATP) in most organisms. The catalysis involves the rotation of a ring of c-subunits, which is driven by the transmembrane electrochemical gradient. This dissertation reports how the eukaryotic c-subunit from spinach chloroplast ATP synthase has successfully been expressed in Escherichia coli and purified in mg quantities by incorporating a unique combination of methods. Expression was accomplished using a codon optimized gene for the c-subunit, and it was expressed as an attachment to the larger, more soluble, native maltose binding protein (MBP-c1). The fusion protein MBP-c1 was purified on an affinity column, and the c1 subunit was subsequently severed by protease cleavage in the presence of detergent. Final purification of the monomeric c1 subunit was accomplished using reversed phase column chromatography with ethanol as an eluent. Circular dichroism spectroscopy data showed clear evidence that the purified c-subunit is folded with the native alpha-helical secondary structure. Recent experiments appear to indicate that this monomeric recombinant c-subunit forms an oligomeric ring that is similar to its native tetradecameric form when reconstituted in liposomes. The F-type ATP synthase c-subunit stoichiometry is currently known to vary from 8 to 15 subunits among different organisms. This has a direct influence on the metabolic requirements of the corresponding organism because each c-subunit binds and transports one H+ across the membrane as the ring makes a complete rotation. The c-ring rotation drives rotation of the gamma-subunit, which in turn drives the synthesis of 3 ATP for every complete rotation. The availability of a recombinantly produced c-ring will lead to new experiments which can be designed to investigate the possible factors that determine the variable c-ring stoichiometry and structure.
ContributorsLawrence, Robert Michael (Author) / Fromme, Petra (Thesis advisor) / Chen, Julian J.L. (Committee member) / Woodbury, Neal W. (Committee member) / Arizona State University (Publisher)
Created2011
Description
In this dissertation I present data gathered from an eleven-month qualitative research study with adolescents living and working on the streets of Lima, Peru. Through the pairing of photovoice with participant observations, this work incorporates distinctive methodological and theoretical viewpoints in order to complicate prevailing understandings of street life. In this dissertation, I examine the identities that children and adolescents on the street develop in context, and the ways in which photography can be a useful tool in understanding identity development among this population. Through a framework integrating theories of identity and identity performance with spatial theories, I outline how identity development among children and adolescents living on the street is directly connected to their relationships with the urban landscape and the outreach organizations that serve them. The organizations and institutions that surround children on the street shape who they are, how they are perceived by society, and how they view and understand themselves in context. It is through the interaction with aid organizations and the urban landscape that a street identity is learned and developed. Furthermore, as organizations, children and adolescents come together within the context of the city, a unique street space is created. I argue that identity and agency are directly tied to this space. I also present the street as a thirdspace of possibility, where children and adolescents are able to act out various aspects of the self that they would be unable to pursue otherwise. Weaved throughout this dissertation are non-traditional writing forms including narrative and critical personal narrative addressing my own experiences conducting this research, my impact on the research context, and how I understand the data gathered.
ContributorsJoanou, Jamie Patrice (Author) / Swadener, Beth B. (Thesis advisor) / Margolis, Eric (Committee member) / Arzubiaga, Angela (Committee member) / Fischman, Gustavo (Committee member) / Arizona State University (Publisher)
Created2011
Description
The Civil Rights Project estimates that Black girls are among the least likely to graduate from high school. More specifically, only about half, or 56%, of freshman Black girls graduate with their class four years later. Beyond the statistics little is known about Black girls who drop out, why they leave school and what happens to them once they are gone. This study is a grounded theory analysis of the stories eight adult Black women told about dropping out of high school with a particular focus on how dropping out affected their lives as workers, mothers and returners to education. There is one conclusion about dropping out and another about Black female identity. First, the women in my study were adolescents during the 1980s, experienced life at the intersection of Blackness, womaness, and poverty and lived in the harsh conditions of a Black American hyperghetto. Using a synthesis between intersectionality and hyperghettoization I found that the women were so determined to improve their economic and personal conditions that they took on occupations that seemed to promise freedom, wealth and safety. Because they were so focused on their new lives, their school attendance suffered as a consequence. In the second conclusion I argued that Black women draw their insights about Black female identity from two competing sources. The two sources are their lived experience and popular controlling images of Black female identity.
ContributorsGriffin, Erica Nicole (Author) / Powers, Jeanne (Thesis advisor) / Fischman, Gustavo (Committee member) / Margolis, Eric (Committee member) / Arizona State University (Publisher)
Created2011
Description
ABSTRACT Early childhood education (ECE) teacher professional development refers to the various modalities of providing new and or additional content knowledge to the teachers who work with children birth to five. The purpose of this study was to examine the effectiveness of an Arizona United Way-administered intervention project designed to provide focused professional development activities to 15 ECE teachers at seven high-need, center-based early care and education settings. Specifically, this study determined if these interventions influenced the teachers to undertake formative career path changes such as college coursework. In addition, the study also sought to understand the views, beliefs, and attitudes of these ECE teachers and if/how their perspectives influenced their educational career paths. Data were gathered through the triangulated use of participants' responses to a survey, face-to-face interviews, and a focus group. Findings demonstrate that the teachers understand that professional development, such as college coursework, can increase a person's knowledge on a given topic or field of study, but that they feel qualified to be a teacher for children birth to five even though 12 of the 15 teachers do not hold an AA/AAS or BA/BS degree in any area of study. Further, the teachers suggested that if they were to earn a degree it would most likely be in another field of study beside education. These responses provide another reason professional development efforts to encourage ECE teachers to seek degrees in the field of education may be failing. If ECE teachers wanted to invest time, energy and funds they would acquire a degree, which provided more financial reward and professional respect.
ContributorsOrtiz, Karen J. (Karen Jean) (Author) / Kelley, Michael F. (Thesis advisor) / Enz, Billie J. (Thesis advisor) / Romero, Mary (Committee member) / Fischman, Gustavo (Committee member) / Arizona State University (Publisher)
Created2011