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Studies on enzymes involved in the biosynthesis of pterin cofactorsBaker, Stephen John January 1997 (has links)
No description available.
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Chemical Transformations Encoded by a Streptomyces coelicolor Gene Cluster with an Unusual GTP CyclohydrolaseSpoonamore, James Edward January 2008 (has links)
Bacterial secondary metabolite biosynthetic pathways are frequently encoded in gene clusters. Genomic sequence information allows the identification of likely biosynthetic clusters based on sequence homology to known proteins. Biochemical characterization of suspected biosynthetic enzymes affords the discovery of pathways which may never be identified by traditional screening approaches. In the work presented here, I, in some cases in collaboration with others, characterize the three intragenomic GTP cyclohydrolase II (GCH II) homologs from Streptomyces coelicolor A3(2) and show that one catalyzes a related but distinct reaction from the other two. The basis for the altered activity is investigated and speaks to the chemical mechanism of not only the unusual enzyme but also to all GCH II enzymes. Further, I investigate two other enzymes found in the same gene cluster as the unusual GCH II. Using biochemical techniques, I show that the product of the unusual GCH II is used as a substrate by a creatinine amidohydrolase homolog. Using structural biology, I show that the third enzyme, a 6-pyruvoyltetrahydropterin synthase (PTPS), can not catalyze the PTPS reaction but is capable of binding a pterin substrate. Finally, I propose that the cluster from S. coelicolor containing the unusual GCH II encodes enzymes for a novel pathway to produce a pterin.
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GTP-Cyclohydrolase function in parasitic nematode developmentBaker, Rachael Helen January 2012 (has links)
Parasitic nematodes of grazing livestock represent an increasing economic and welfare problem for British agriculture. By investigating specific life-cycle stages of these parasites, it may be possible to identify key molecules or pathways that are required for the survival of the worms, and thus exploit these for future control strategies. It has been shown previously that the third larval stages (L3) of the ovine parasitic nematode Teladorsagia circumcincta produce high levels of transcript for the enzyme GTP-Cyclohydrolase relative to later developmental stages. As the ratelimiting factor in the production of tetrahydrobiopterin, GTP-Cyclohydrolase is required for a number of different biochemical pathways, including those involved in the production of serotonin and melanin. As the L3 do not feed, it can be hypothesised that, if finite resources are being used in the production of transcript encoding this enzyme, then it may be important for survival. In this thesis, a number of approaches were taken to explore the function of GTPCyclohydrolase in the life-cycle development of T. circumcincta. The closely related parasite, Dictyocaulus viviparus, was used as a model organism to explore the role of GTP-Cyclohydrolase and serotonin production with regards to larval arrest, or hypobiosis. This process occurs readily under experimental conditions in D. viviparus, which is not possible with T. circumcincta. Quantitative PCR was used to examine GTP-Cyclohydrolase transcript levels in two different strains of D. viviparus, one that enters larval arrest when exposed to cold conditions and one that does not. No differences were observed between the two strains suggesting that GTP-Cyclohydrolase was unlikely to be involved in hypobiosis. The model nematode, Caenorhabditis elegans, was used to perform functional complementation experiments to assess the role of GTP-Cyclohydrolase in the cuticle, as it has been shown previously that C. elegans GTP-Cyclohydrolase mutants have a ‘leaky cuticle’ and are killed by lower doses of anthelmintics and bleach than the wild-type worms. The T. circumcincta gene for GTP-Cyclohydrolase was able to restore cuticular integrity of C. elegans GTP-Cyclohydrolase-deletion mutants, suggesting that the role played by the protein in both species is similar. In vitro inhibition experiments using a chemical inhibitor of GTP-Cyclohydrolase showed that T. circumcincta larval development was disrupted in the presence of the inhibitor. It was also shown that T. circumcincta L3 that were exposed to sunlight produced melanin, suggesting that the levels of GTP-Cyclohydrolase observed in the preparasitic stages of T. circumcincta may be required for the synthesis of melanin. Together, these data suggest that GTP-Cyclohydrolase is required by the preparasitic stages to survive on pasture. Ultraviolet radiation has been shown previously to be harmful to T. circumcincta L3, so if the melanin production provides protection from this, then it would be crucial for the survival of the pre-parasitic stages.
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Untersuchung von Strukturfunktionsbeziehungen bei Enzymen der Tetrahydrobiopterin- und RiboflavinbiosyntheseSchiffmann, Susanne. January 2002 (has links) (PDF)
München, Techn. Univ., Diss., 2002.
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Metabolic engineering of the pterin branch of folate synthesis by over-expression of a GTP cyclohydrolase I in peanutJuba, Nicole Czarina 11 November 2011 (has links)
Folate, also known as vitamin B9, is an essential dietary vitamin that provides the donor group for one carbon transfer reactions. Deficiency in folate is associated with neural tube birth defects (NTDs), cancer, cardiovascular disease, and anemia. In the US enriched food products including bread, pasta, and cereal are fortified with folic acid, the synthetic analog of folate. While effective in reducing NTDs, this practice is costly and not economically practical in developing countries. Folate biofortification, increasing the natural folate level in foods by metabolic engineering, has been proposed as a sustainable alternative to food fortification with folic acid. To increase folate levels in peanut seed, GTP cyclohydrolase I from Arabidopsis thaliana (AtGCHI) was introduced into peanut by biolistic transformation. Plant transformation vectors were constructed using publicly available or licensable vector components to avoid intellectual property restrictions that hinder commercialization. Thirteen peanut cultivars were evaluated for transformation efficiencies and regeneration potential. Expression levels of the AtGCHI transgene were determined by quantitative real-time PCR. The endogenous peanut GCHI (AhGCHI) was isolated and sequenced. Studies were conducted to test whether heterologous over-expression of AtGCHI altered expression of the endogenous AhGCHI. Seed-specific expression of AtGCHI does not affect AhGCHI transcript accumulation. For validation of the proposed folate biofortification strategy, vitamin quantification will be required. A liquid chromatography tandem mass spectrometry (LC/MS/MS) method was developed to identify and quantify the different forms of folate. However, additional work will be needed to determine sensitivity of the instrument, to optimize vitamin extraction, and to increase sufficient seed for vitamin extraction and analysis. Peanut products derived from folate biofortified peanut kernels will have a niche market in the United States, but there is a larger global implication as a mechanism for sustainable delivery of essential vitamins to populations that can not adopt synthetic vitamin supplementation/fortification. Successful demonstration of increased folate in peanut will result in better vitamin availability for populationssonsuming peanut based foods as a dietary staple. / Ph. D.
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