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Evolution of copper-containing nitrite reductaseMacPherson, Iain 05 1900 (has links)
Copper-containing nitrite reductase (NiR) is a homotrimer of two cupredoxin domains and catalyzes the single electron reduction of NO2- to NO during dissimilatory denitrification. To investigate the evolution of NiR, methods of mutagenic library generation and high-throughput variant screening from E. coli colonies were developed. These methods allow for facile screening of 105 mutants for folding efficiency or substrate specificity. Initial proof of principle studies yielded several variants that oxidized the artificial substrate ο-dianisidine up to 8 times faster than wild type NiR, suggesting that this methodology has the potential to engineer NiR to acquire other reductase functions.
A crystal structure was solved for a putative multicopper oxidase (MCO) and NiR homologue from Arthrobacter sp. (AMMCO) to 1.8 Å resolution. The overall folds of AMMCO and NiR are very similar (r.m.s.d. of 2.0 Å over 250 Cα atoms); Like NiR, AMMCO is a trimer with type-1 Cu sites in the N-terminal domain of each monomer; however, the active site of AMMCO contains trinuclear Cu site characteristic of MCOs instead of a the mononuclear type-2 Cu site found in NiR. Detailed structural analysis supports the theory that two-domain MCOs similar to AMMCO were intermediaries in the evolution of NiR and the more common three-domain MCOs. The physiological function of AMMCO remains uncertain, but genomic, crystallographic and functional analysis suggests that the enzyme is involved in metal regulation.
Considering the extensive similarity between AMMCO and NiR, particularly at the active site, engineering a trinuclear cluster into NiR appears feasible with a modest number of alterations to the polypeptide chain. With the aid of my newly developed high-throughput screening technique and site-directed mutagenesis, the mononuclear NiR active site was remodelled into a trinuclear Cu site similar to that of MCO. A crystal structure of this variant was solved to 2.0 Å and the presence of three copper atoms at the engineered cluster was confirmed by Cu-edge anomalous diffraction data. Although the trinuclear copper cluster is present and catalyzes the reduction of oxygen, achieving rates of catalysis seen in native MCOs has proven more difficult. With the framework provided, further engineering NiR into a robust MCO is likely to provide further insights into the structural basis of oxygen reduction by trinuclear copper sites.
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The Search for Genetic Structure and Patterns in Vietnamese FrogsNgo, Andre 01 August 2008 (has links)
Vietnam has the greatest biodiversity of any country in Indochina. This diversity may be due to its topographically complex nature, with hills and mountains, drained by several independent river systems, covering three quarters of its area. Topographic complexity has undoubtedly had profound effects on the flora and fauna of the region. Recent surveys have uncovered several cryptic species in what were previously considered single widespread species. These discoveries have led some researchers to propose that widespread forest species do not, in fact, exist in Southeast Asia. To test these hypotheses, I examined patterns of mitochondrial phylogeny in several groups of frogs, both at and below the species level. Additionally, these analyses helped clarify the otherwise chaotic picture of anuran taxonomy and systematics. The stream–tied waterfall frogs of the genera Amolops and Odorrana were examined, the monophyly of the ranid subfamily Amolopinae was rejected, and taxonomic adjustments were made. The phylogeny of the Vietnamese narrow–mouthed frogs of the genus Microhyla was recovered and the current taxonomy examined. Patterns of maternal dispersal and genetic differentiation in mitochondrial DNA were further examined within Microhyla heymonsi, revealing geographic structuring and the existence of two sympatric lineages. Lastly, frogs of the Polypedates leucomystax complex were examined and two major, largely sympatric lineages recovered. Within these groups, 11 separate mitochondrial lineages identified. These represented separate species on the basis of advertisement call and allozyme evidence. The relationship of genetic differentiation and river systems was also investigated and common patterns among the different groups were explored. Clear genetic breaks occurred across both the Red River and the Annamite Mountain range, though most common patterns were groupings of populations along river drainages. While several cryptic species were identified, widespread groups likely representing single species still exist, and a phylogenetic component to broad distribution were noted.
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Modeling EvolutionEarnshaw-Whyte, Eugene 04 March 2013 (has links)
Evolution by natural selection began as a biological concept, but since Darwin it has been recognized to have broader application than biology. Applying evolutionary ideas beyond biology requires that the principles of evolution by natural selection be abstracted and generalized from the biological case. The received view of evolution by natural selection in biology is itself seriously flawed, which understandably renders the project of abstracting it and applying it elsewhere challenging. This thesis develops a generalized account of models of evolution by natural selection which is used to resolve various outstanding issues in the philosophy of biology. This also clarifies the methods and prospects of applying evolution by natural selection to non-biological domains. It does so by analyzing models of evolution both within biology and outside it, relying in particular on the contrast provided by models of firm competition in evolutionary economics. This analysis highlights those aspects of the classical view which must be abandoned or revised, and leads to the development of a neo-dynamical model of evolution, which is developed, explained, defended, and applied to problems in evolutionary biology and multi-level selection theory.
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Evolution of Chromatin Modification MachineryOn, Tuan 13 January 2011 (has links)
This thesis explores chromatin modification (CM) as a biological system and uses known CM factors in four model organisms; yeast, worm, fly, and human to explore how CM factors have consistently evolved across a diverse spectrum of 111 organisms by using the InParanoid homology algorithm. Using InParanoid, phylogenetic profiles are constructed for each model organism to highlight evolutionary trajectories and which CM factors are lost, expanded, and are specific to some lineages. Phylogenetic tree construction demonstrates that peripheral subunits of CM complexes evolve independently. Accurate mapping of domains to CM factors and their homologs reveals that the architecture of domains is very well conserved, with only one potential case of a domain swap. Homology, domain architecture, and protein-protein interaction is then combined to illustrate an interolog example and potential interaction candidates. The techniques highlighted in this thesis represent a generic and powerful approach to analyzing any biological system of interest.
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Transcriptome Assembly and Molecular Evolutionary Analysis of Sex-biased Genes in Caenorhabditis Species 9 and Caenorhabditis Species 5Rajagopalan, Deepthi 26 November 2012 (has links)
Differential gene expression between sexes is the main contributor of the morphological and behavioral differences observed between them. Studying the signatures of these differences at the genetic level will help us understand the forces acting on them. The existence of androdioecious and gonochoristic species in the genus Caenorhabditis makes it suitable for sex-biased gene expression studies. In this thesis, I have assembled the transcriptome of C. sp. 9 and C. sp. 5 using de novo and reference-based techniques. Evolutionary analysis of the assembled contigs showed that genes with male-biased expression evolve faster than those with a female bias, as observed in other taxa. Furthermore, I found a positive correlation between gene expression and codon usage bias.
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Evolution of Chromatin Modification MachineryOn, Tuan 13 January 2011 (has links)
This thesis explores chromatin modification (CM) as a biological system and uses known CM factors in four model organisms; yeast, worm, fly, and human to explore how CM factors have consistently evolved across a diverse spectrum of 111 organisms by using the InParanoid homology algorithm. Using InParanoid, phylogenetic profiles are constructed for each model organism to highlight evolutionary trajectories and which CM factors are lost, expanded, and are specific to some lineages. Phylogenetic tree construction demonstrates that peripheral subunits of CM complexes evolve independently. Accurate mapping of domains to CM factors and their homologs reveals that the architecture of domains is very well conserved, with only one potential case of a domain swap. Homology, domain architecture, and protein-protein interaction is then combined to illustrate an interolog example and potential interaction candidates. The techniques highlighted in this thesis represent a generic and powerful approach to analyzing any biological system of interest.
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The Search for Genetic Structure and Patterns in Vietnamese FrogsNgo, Andre 01 August 2008 (has links)
Vietnam has the greatest biodiversity of any country in Indochina. This diversity may be due to its topographically complex nature, with hills and mountains, drained by several independent river systems, covering three quarters of its area. Topographic complexity has undoubtedly had profound effects on the flora and fauna of the region. Recent surveys have uncovered several cryptic species in what were previously considered single widespread species. These discoveries have led some researchers to propose that widespread forest species do not, in fact, exist in Southeast Asia. To test these hypotheses, I examined patterns of mitochondrial phylogeny in several groups of frogs, both at and below the species level. Additionally, these analyses helped clarify the otherwise chaotic picture of anuran taxonomy and systematics. The stream–tied waterfall frogs of the genera Amolops and Odorrana were examined, the monophyly of the ranid subfamily Amolopinae was rejected, and taxonomic adjustments were made. The phylogeny of the Vietnamese narrow–mouthed frogs of the genus Microhyla was recovered and the current taxonomy examined. Patterns of maternal dispersal and genetic differentiation in mitochondrial DNA were further examined within Microhyla heymonsi, revealing geographic structuring and the existence of two sympatric lineages. Lastly, frogs of the Polypedates leucomystax complex were examined and two major, largely sympatric lineages recovered. Within these groups, 11 separate mitochondrial lineages identified. These represented separate species on the basis of advertisement call and allozyme evidence. The relationship of genetic differentiation and river systems was also investigated and common patterns among the different groups were explored. Clear genetic breaks occurred across both the Red River and the Annamite Mountain range, though most common patterns were groupings of populations along river drainages. While several cryptic species were identified, widespread groups likely representing single species still exist, and a phylogenetic component to broad distribution were noted.
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Understanding pathogen selection pressures at the within- and between-host levelsBall, Colleen 11 1900 (has links)
Many infectious pathogens, and in particular viruses, have an
extremely high rate of mutation. This can lead to rapid evolution
driven by selection pressures operating at both the within- and
between-host levels, as strains compete for resources within their
chosen host while also competing to effectively transmit to new
hosts. In the case of chronic viral infections, such as the human
immunodeficiency virus (HIV) or hepatitis C, substantial viral
evolution may take place within a single infected host. The fitness
of a pathogen has been studied at the between-host level and at the
within-host level, but linking the two levels of selection pressure
is a difficult problem that has yet to be studied satisfactorily.
We modify a simple model describing the within host dynamics of HIV
infection by including multiple pathogen strains with different
properties and allowing these strains to mutate. Within the host we
observe different strategies for pathogen success during different
stages of infection, which often leads to different strains
predominating within the host over the course of infection. We then
embed our within-host model into a Monte Carlo simulation that
models the interactions between infected individuals. This approach
allows us to combine selective pressure at the within-host level
with pressures at the between-host level and helps us to predict
which strains are most likely to be present within the population.
We show that under our model assumptions the co-existence of
multiple strains is possible and we explore the factors leading to
the success of a pathogen.
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189 |
Understanding pathogen selection pressures at the within- and between-host levelsBall, Colleen 11 1900 (has links)
Many infectious pathogens, and in particular viruses, have an
extremely high rate of mutation. This can lead to rapid evolution
driven by selection pressures operating at both the within- and
between-host levels, as strains compete for resources within their
chosen host while also competing to effectively transmit to new
hosts. In the case of chronic viral infections, such as the human
immunodeficiency virus (HIV) or hepatitis C, substantial viral
evolution may take place within a single infected host. The fitness
of a pathogen has been studied at the between-host level and at the
within-host level, but linking the two levels of selection pressure
is a difficult problem that has yet to be studied satisfactorily.
We modify a simple model describing the within host dynamics of HIV
infection by including multiple pathogen strains with different
properties and allowing these strains to mutate. Within the host we
observe different strategies for pathogen success during different
stages of infection, which often leads to different strains
predominating within the host over the course of infection. We then
embed our within-host model into a Monte Carlo simulation that
models the interactions between infected individuals. This approach
allows us to combine selective pressure at the within-host level
with pressures at the between-host level and helps us to predict
which strains are most likely to be present within the population.
We show that under our model assumptions the co-existence of
multiple strains is possible and we explore the factors leading to
the success of a pathogen.
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Intron retention and recognition in the microsporidian encephalitozoon cuniculiLee, Renny 11 1900 (has links)
Microsporidia are unicellular fungi that are intracellular parasites of animals, including
humans. They are both complex and simple, armed with a sophisticated infection
apparatus and possessing the smallest eukaryotic nuclear genomes. The microsporidian
Encephalitozoon cuniculi has a genome size of 2.9 Mb, which is smaller than many
bacterial genomes. Genome reduction and compaction in size, content, and form has been
interpreted as an adaptation to parasitism. One of the effects of genome size reduction
concerns intron evolution — E. cuniculi has retained only a few extremely short
spliceosomal introns.
This thesis examines the splicing of introns in the spore stage. The introns were retained
in spores, suggesting life-stage specific splicing and splicing inhibition. How the short
introns are recognized was also examined. Unique splicing signal motifs were predicted,
and were used to find additional introns. The intron density was doubled for this species,
and I also obtained data that counter current views about intron evolution in compacted
genomes with low intron densities. I also predict that E. cuniculi introns are recognized
in a unique way by the spliceosome.
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