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Thesis (Ph. D.)--University of Hawaii at Manoa, 2005. / Includes bibliographical references (leaves 101-111). / Also available by subscription via World Wide Web / xii, 111 leaves, bound ill. 29 cm

Thesis (Ph. D.)--University of Hawaii at Manoa, 2005. / Includes bibliographical references (leaves 101-111).

Thesis (Ph. D.)--University of Wisconsin--Madison, 1970. / Typescript. Vita. Description based on print version record. Includes bibliographical references (leaves 98-106).

Abstract Type XII minicollagen chain association was studied using baculovirus-directed insect cell expression. Since insect cells contain low endogenous prolyl 4-hydroxylase activity, the mechanism of the effect of prolyl hydroxylation on trimer formation in this collagen could be studied directly by adding recombinant baculoviruses directing the synthesis of prolyl 4-hydroxylase. Prolyl 4-hydroxylase was shown to be involved in the trimeric assembly process of type XII collagen through its α subunit, and thus through its hydroxylase activity. The transmembrane protein type XIII collagen was also characterized by means of insect cell expression, for which purpose new antibodies against its non-collagenous domains NC2 and NC4 were generated, together with a pan-collagen antibody against collagenous sequences. Type XIII collagen α chains were found to form disulphide-bonded homotrimers, and this was enhanced by prolyl 4-hydroxylation. Analysis of the disulphide-bonding pattern of the eight cysteine residues of the α1(XIII) chains revealed that some of the cysteines in the NC1 domain, and possibly the cysteines at the junction of the COL1 and NC2 domains, are interchain-linked, while the cysteines in the NC4 domain are intrachain-linked. The three collagenous domains of type XIII collagen were shown to be in triple-helical conformation and have different thermal stabilities, i.e. 38±C for the COL1 domain, 49±C for COL2 and 40±C for COL3. Furthermore, it was shown that type XIII collagen is oriented in the plasma membrane of insect cells so that its non-collagenous N-terminus is intracellular and its mostly collagenous C-terminus is extracellular. Type XIII collagen was also found to be cleaved into the insect cell culture medium by a furin-like protease. The expression of various type XIII collagen deletion variants suggested that chain recognition and the association of type XIII collagen α chains into trimers occur in the N-terminal portion of this molecule. An internal in-frame deletion of residues 63-83 immediately adjacent to the transmembrane domain indicated that this short ectodomain sequence is necessary for the formation of disulphide-bonded trimers. Since a sequence homologous with these deleted residues was also found at the same plasmamembrane-adjacent location in other collagenous transmembrane proteins, this points to common features in their chain association.

published_or_final_version / Chemistry / Doctoral / Doctor of Philosophy

This abstract sent to supervisor 5/4/12 Tyrosine hydroxylase is the enzyme which controls the ratelimiting step in the synthesis of noradrenaline. In order to discover whether the activity of tyrosine hydroxylase might serve as an indicator of noradrenergic function in the brain, two preparations for the direct measurement of TH activity in rat brain regions by a tritium-release method were employed: synaptosomal suspensions prepared from pinchedoff nerve terminals, and partially solubilised enzyme preparations prepared from frozen homogenates or synaptosomal suspensions and assayed at saturating concentrations of cofactor and tyrosine. There was evidence of an increase in tyrosine hydroxylase activity in hippocampal synaptosomes of rats killed immediately after a mild electrical footshock. Activation of synaptosomal enzyme activity was also found after single doses of clonidine and parachloroamphetamine, and after repeated handling; and single doses of morphine and of yohimbine appeared to lower tyrosine hydroxylase activity. Repeated administration of drugs such as clonidine, desipramine and 2-deoxyglucose, however, did not affect tyrosine hydroxylation rate. A preliminary finding , suggesting differences in synaptosomal tyrosine hydroxylase activity related to experience with different reinforcement schedules (continuous reward vs. partial reward) in a runway experiment, was not substantiated in later experiments; nor was there any difference between the synaptosomal tyrosine hydroxylase activity of naive controls and rats given repeated daily shocks for a week. The saturated TH assay performed on solubilised enzyme was, as predicted, unresponsive to the short term stimulation effects detected with the synaptosomal assay. However, other changes, such as a reduced maximal hydroxylation rate after repeated desipramine administration, and an increased rate several weeks after a course of electrical stimulation of the septal area, were established with the saturated assay. Although the changes in stimulated rats were associated with increased behavioural tolerance to stress, e.g. resistance to extinction of a running response in a runway, other experiments in which the behavioural stress-tolerance was induced by behavioural methods alone showed no accompanying changes in TH activity. Measures of synaptosomal and saturated soluble TH activity appear to constitute independent indicators of noradrenergic function. It seems that synaptosomal tyrosine hydroxylase activity is not, as anticipated, controlled by the firing rate of locus coeruleus neurones; but it may be subject to local regulation in noradrenergic terminals. The results are discussed in the context of theoretical aspects of the regulation of noradrenaline synthesis in the brain, and the mechanisms underlying physiological responses to stress and behavioural tolerance to stress.

Thesis (Ph. D.)--University of Adelaide, Dept. of Clinical and Experimental Pharmacology, 1989. / Includes bibliographical references (leaves 162-174).

Th. univ.--Sci. pétrolières, spécialité Chimie--Paris VI et Ecole nationale supérieure du pétrole et des moteurs, 1987. / 1988 d'après la déclaration de dépôt légal. Bibliogr. p. 181-188.