Thesis (Doctoral)--Universität Hamburg, 1982.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1966. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

(2S)- and (2R)-Serine O-sulphate have been synthesised and shown to inactivate glutamate decarboxylase (GAD) from E. Coli. Novel methodology was developed to enable the stereospecific synthesis of (2S) and (2R)-deuteriated serine in order to probe the mechanism of inactivation. The rates of inactivation of glutamate decarboxylase by (2S)-, (2S)-[2-2H]-, (2R)- and (2R)-[2-2H]-serine O-sulphate have been measured for each of the isotopomers at a range of concentrations. From the data obtained the deuterium isotope effects were determined for each enantiomer. The inactivation by the (2S)-enantiomer was shown to involve C-H bond cleavage while inactivation by the (2R)-isomer involves C-decarboxylation. Both processes were shown to occur on the 4'-re-face of the coenzyme, the opposite face to that utilised in the physiological decarboxylation reaction. The methodology developed for the synthesis of the deuteriated serines involved the regiospecific introduction of deuterium to the C-6 centre of (3R)- and (3S)-2,5- dimethoxy-3-isopropyl-3,6-dihydropyrazine. Schollkopf chemistry was then exploited for the stereospecific alkylation at C-6 of the dihydropyrazines. This chemistry was versatile and enabled the synthesis of other deuteriated amino acids. For example (2S)-[2-2H]-phenylalanine, (2S)-[2-2H]-allylglycine and (2S)-[2-2H]-aspartic acid were synthesised using this chemistry. The decarboxylation of 2-aminomalonic acid by cytosolic serine hydroxymethyltransferase (SHMT) was studied. Contrary to previous reports, the reaction was found to be stereospecific and the newly introduced hydrogen was shown to occupy the 2-pro-S position of the glycine product.

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A major pathway of tyramine and dopamine synthesis in insects is through the decarboxylation of tyrosine and DOPA, respectively. Although tyrosine decarboxylase (TDC) has been mentioned in some reports, it has never been critically analyzed. The high sequence identity shared by tyrosine decarboxylase and DOPA decarboxylase in insects, and the similar structures of the substrates, tyrosine and DOPA, raise the possibility that both tyrosine decarboxylase and DOPA decarboxylase (DDC) have activities to tyrosine and DOPA. In this study, after tyrosine decarboxylase and DOPA decarboxylase enzymes of Anopheles gambiae were expressed, their substrate specificities and biochemical properties were critically analyzed. My results provide clear biochemical evidence establishing that the mosquito tyrosine decarboxylase functions primarily on the production of tyramine with low activity to DOPA. In contrast, mosquito DOPA decarboxylase is highly specific to DOPA with essentially no activity to tyrosine. / Master of Science in Life Sciences

ODC activity of the altered proteins was measured and compared to that of the full length 461 amino acid containing ODC. Mouse ODC cDNA sequences were deleted from either 5' or 3' ends using exonuclease III and Mung Bean nuclease treatments. An internal deletion was obtained by Hinc II and Bcl I restriction endonuclease digestion of the full length ODC cDNA. Capped mRNAs were synthesized in vitro using the resulting deleted DNA as templates, and the protein was translated in vitro. The results indicate that the protein in which translation initiates at internal AUG start codons does not have any activity. The protein with 39 amino acids deleted from carboxy-terminus maintains 12% of the activity, while deletion of greater than 79 amino acids have no activity. An internal deletion from amino acid 290 to 331 and which may contain the suspected ornithine binding site has no activity. These results suggest that the entire amino acid sequence of mouse ODC is required for full activity of the enzyme.

Bibliography: leaves 255-280. In vivo and in vitro investigations of the hypothesis that polyamines and their synthetic enzymes have a role in regulation of cellular and molecular processes in the follicle. The activity of ornithine decarboxylase (ODC), the rate limiting polyamine biosynthetic enzyme, is demonstrated in wool follicle homogenates.

The objectives of this study were to identify genetic variants, develop the respective haplotypes (combination of alleles) and investigate the association of identified variants with economically important traits in two candidate genes. The first gene was Phosphoenolpyruvate carboxykinase (PEPCK) which is a key regulatory enzyme of gluconeogenesis. The second candidate gene, Ornithine decarboxylase (ODC), is a rate-limiting enzyme in polyamine biosynthesis. It has a significant role in DNA synthesis and cell proliferation. We first analyzed the genetic variability of PEPCK-C, the gene which codes for the cytosolic form of PEPCK. A 3792 by segment of 5'-region of the PEPCK-C gene (pos. -1723 to 2069) was sequenced in 32 White Leghorn chickens (a total of 64 genomes). A total of 19 single nucleotide polymorphisms (SNPs) were identified. We then analyzed the genetic variability of ODC. A 5 kb sequence of 3' end of the gene was sequenced in 20 White Leghorn chickens (a total of 40 genomes). A total of 63 variant sites were identified. The rate of insertion/deletion in ODC was 16%, whereas neither deletions nor insertions were present in PEPCK-C. Gene trees were constructed for both genes assuming maximal parsimony. This led to the delineation of 6 haplotypes in PEPCK-C. Two of the SNPs coincided with RFLP detectable by the restriction enzymes AciI and BstEII, respectively. Three haplotypes in ODC were defined. In the next step, White Leghorn chickens from a non-selected closed population were typed for these two PEPCK-C RFLP. The two RFLP gave rise to three alleles (or haplotype classes), which in turn defined six genotypes. A comparison of genotypes revealed significant differences in feed efficiency (FE) and residual feed consumption (RFC). There was significant interaction between PEPCK-C genotypes and mitochondrial PEPCK (PEPCK-M) genotypes defined by an RFLP. The latter enzyme catalyzes the same reaction, but is located in the matrix of t

Includes bibliographical references (leaves 255-280) / xv, 280 leaves, [24] leaves of plates : ill. (some col.) ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / In vivo and in vitro investigations of the hypothesis that polyamines and their synthetic enzymes have a role in regulation of cellular and molecular processes in the follicle. The activity of ornithine decarboxylase (ODC), the rate limiting polyamine biosynthetic enzyme, is demonstrated in wool follicle homogenates. / Thesis (Ph.D.)--University of Adelaide, Dept. of Animal Science, 1995