Matching Items (375)
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
The purpose of this project was to commission, perform, and discuss a new work for an instrument pairing not often utilized, oboe and percussion. The composer, Alyssa Morris, was selected in June 2009. Her work, titled Forecast, was completed in October of 2009 and premiered in February of 2010, as part of a program showcasing music for oboe and percussion. Included in this document is a detailed biography of the composer, a description of the four movements of Forecast, performance notes for each movement, a diagram for stage set-up, the full score, the program from the premiere performance with biographies of all the performers involved, and both a live recording and MIDI sound file. The performance notes discuss issues that arose during preparation for the premiere and should help avoid potential pitfalls. TrevCo Music, publisher of the work, graciously allowed inclusion of the full score. This score is solely for use in this document; please visit the publisher's website for purchasing information. The commission and documentation of this composition are intended to add to the repertoire for oboe in an unusual instrument pairing and to encourage further exploration of such combinations.
ContributorsCreamer, Caryn (Author) / Schuring, Martin (Thesis advisor) / Hill, Gary (Committee member) / Holbrook, Amy (Committee member) / Micklich, Albie (Committee member) / Spring, Robert (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
During the twentieth-century, the dual influence of nationalism and modernism in the eclectic music from Latin America promoted an idiosyncratic style which naturally combined traditional themes, popular genres and secular music. The saxophone, commonly used as a popular instrument, started to develop a prominent role in Latin American classical music beginning in 1970. The lack of exposure and distribution of the Latin American repertoire has created a general perception that composers are not interested in the instrument, and that Latin American repertoire for classical saxophone is minimal. However, there are more than 1100 works originally written for saxophone in the region, and the amount continues to grow. This Modern Latin American Repertoire for Classical Saxophone: Recording Project and Performance Guide document establishes and exhibits seven works by seven representative Latin American composers.The recording includes works by Carlos Gonzalo Guzman (Colombia), Ricardo Tacuchian (Brazil), Roque Cordero (Panama), Luis Naón (Argentina), Andrés Alén-Rodriguez (Cuba), Alejandro César Morales (Mexico) and Jose-Luis Maúrtua (Peru), featuring a range of works for solo alto saxophone to alto saxophone with piano, alto saxophone with vibraphone, and tenor saxophone with electronic tape; thus forming an important selection of Latin American repertoire. Complete recorded performances of all seven pieces are supplemented by biographical, historical, and performance practice suggestions. The result is a written and audio guide to some of the most important pieces composed for classical saxophone in Latin America, with an emphasis on fostering interest in, and research into, composers who have contributed in the development and creation of the instrument in Latin America.
ContributorsOcampo Cardona, Javier Andrés (Author) / McAllister, Timothy (Thesis advisor) / Spring, Robert (Committee member) / Hill, Gary (Committee member) / Pilafian, Sam (Committee member) / Rogers, Rodney (Committee member) / Gardner, Joshua (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
Finger motion and hand posture of six professional clarinetists (defined by entrance into or completion of a doctorate of musical arts degree in clarinet performance) were recorded using a pair of CyberGloves® in Arizona State University's Center for Cognitive Ubiquitous Computing Laboratory. Performance tasks included performing a slurred three-octave chromatic scale in sixteenth notes, at sixty quarter-note beats per minute, three times, with a metronome and a short pause between repetitions, and forming three pedagogical hand postures. Following the CyberGloves® tasks, each subject completed a questionnaire about equipment, playing history, practice routines, health practices, and hand usage during computer and sports activities. CyberGlove® data were analyzed to find average hand/finger postures and differences for each pitch across subjects, subject variance in the performance task and differences in ascending and descending postures of the chromatic scale. The data were also analyzed to describe generalized finger posture characteristics based on hand size, whether right hand thumb position affects finger flexion, and whether professional clarinetists use similar finger/hand postures when performing on clarinet, holding a tennis ball, allowing hands to hang freely by the sides, or form a "C" shape. The findings of this study suggest an individual approach based on hand size is necessary for teaching clarinet hand posture.
ContributorsHarger, Stefanie (Author) / Spring, Robert (Thesis advisor) / Hill, Gary (Committee member) / Koonce, Frank (Committee member) / Norton, Kay (Committee member) / Stauffer, Sandy (Committee member) / Arizona State University (Publisher)
Created2011
Description
Owen Middleton (b. 1941) enjoys an established and growing reputation as a composer of classical guitar music, but his works for piano are comparatively little known. The close investigation offered here of Middleton's works for piano reveals the same impressive craftsmanship, compelling character, and innovative spirit found in his works for guitar. Indeed, the only significant thing Middleton's piano music currently lacks is the well-deserved attention of professional players and a wider audience. Middleton's piano music needs to be heard, not just discussed, so one of this document's purposes is to provide a recorded sample of his piano works. While the overall repertoire for solo piano is vast, and new works become established in that repertoire with increasing difficulty, Middleton's piano works have a significant potential to find their way into the concert hall as well as the private teaching studio. His solo piano music is highly effective, well suited to the instrument, and, perhaps most importantly, fresh sounding and truly original. His pedagogical works are of equal value. Middleton's piano music offers something for everyone: there one finds daring virtuosity, effusions of passion, intellectual force, colorful imagery, poetry, humor, and even a degree of idiomatic innovation. This study aims to reveal key aspects of the composer's musical style, especially his style of piano writing, and to provide pianists with helpful analytical, technical, and interpretive insights. These descriptions of the music are supported with recorded examples, selected from the works for solo piano written between 1962 and 1993: Sonata for Piano, Childhood Scenes, Katie's Collection, and Toccata for Piano. The complete scores of the recorded works are included in the appendix. A chapter briefly describing the piano pieces since 1993 concludes the study and invites the reader to further investigations of this unique and important body of work.
ContributorsMoreau, Barton Andrew (Author) / Hamilton, Robert (Thesis advisor) / Holbrook, Amy (Committee member) / Campbell, Andrew (Committee member) / Spring, Robert (Committee member) / Gardner, Joshua (Committee member) / Arizona State University (Publisher)
Created2011
Description
As clarinet compositions created by Taiwanese composers have not been widely discussed and published in academia, this research paper examines three unaccompanied clarinet works by three Taiwanese composers: Ballade by Hsiao-Wen Tseng, Chin Thoughts III by Ling-Huei Tsai, and Pointe de Flame by Chia-Lin Pan, all commissioned by the author in 2007. This research also includes a compact disc with recordings of these works, aiming to document the creativity of Taiwanese composers. This research paper examines these three commissioned works by analyzing their overall musical styles, notations, formal structures, rhythmical and melodic materials, unconventional clarinet techniques as well as the influence of Chinese musical elements. The analysis reveals the distinctive characteristic of each piece. Moreover, the author provides composers' insights and performance guides to help interested readers practice these pieces. To further understand how the composers create these pieces by drawing upon different life experiences, the paper also includes information about their backgrounds, program notes, lists of compositions, and music examples for reference. The author found that collaborating with these composers helped to establish a closer composer-performer relationship in interpreting the music. It is hoped that this compact disc recording will help make Taiwanese composers' clarinet works more accessible to a wider audience. Moreover, this research paper hopes to generate more interest in performing and appreciating music composed by Taiwanese composers.
ContributorsChuang, Yenting (Author) / Spring, Robert (Thesis advisor) / Schuring, Martin (Committee member) / Campbell, Andrew (Committee member) / Jiang, Danwen (Committee member) / Hackbarth, Glenn (Committee member) / Arizona State University (Publisher)
Created2011