Thesis (Ph. D.)--University of Florida, 2004. / Title from title page of source document. Document formatted into pages; contains 79 pages. Includes vita. Includes bibliographical references.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1970. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

Studies of the binding of coenzyme analogs to yeast 6-phosphogluconate dehydrogenase indicate that NADP binding to the enzyme results from selective interactions between regions of the coenzyme binding site and portions of the NADP molecule. These studies suggested the existence of coenzyme binding site regions which selectively interact with the adenosine, 2'-phosphate, and pyrophosphate moieties of NADP. The importance of the 2'-phosphate to coenzyme binding was indicated by enhanced binding of adenosine derivatives possessing this moiety when compared to adenosine derivatives not phosphorylated at this position. The better binding of the 2'-phosphorylated derivatives became more pronounced with increasing resemblance of the derivative to the NADP molecule, and NAD was not inhibitory up to 70 roM. These results substantiate the concept that interaction of the enzyme with the d2'-phosphate is a key factor in the specificity of yeast 6-phosphogluconate dehydrogenase for NADP. Structural analogs of the pyridinium portion of the NADP molecule, Nl-alkylnicotinamide chlorides, did not inhibit yeast 6-phosphogluconate dehydrogenase at concentrations normally required for selective interactions with dehydrogenases; however, enzyme activity was decreased at micellar concentrations of Nl-dodecylnicotinamide chloride. Investigations of the role and environment of the essential sulfhydryl group of this enzyme were also performed. N-alkylmaleimides (N-methyl - N-hexyl, inclusive) were shown to inactivate the enzyme, but without a chainlength effect. In the presence of 6-phosphogluconate, the enzyme was protected from N-ethylmaleimide inactivation and this protection was enhanced by the addition of NADPH or AADP. / Master of Science

Thesis (Ph. D.)--University of Oregon, 2000. / Typescript. Includes vita and abstract. Includes bibliographical references (leaves 56-59). Also available for download via the World Wide Web; free to University of Oregon users.

Thesis (B. Sc. Hons)--University of Adelaide, 1984. / Includes bibliographical references (leaves [86-88]).

Thesis (Ph. D.)--University of Oregon, 2007. / Typescript. Includes vita and abstract. Includes bibliographical references (leaves 94-100). Also available for download via the World Wide Web; free to University of Oregon users.

Thesis (Ph. D.)--University of California, San Diego, and San Diego State University, 2004. / Vita. Includes bibliographical references.

Thesis (Ph. D.)--University of California, San Diego and San Diego State University, 2005. / Vita. Includes bibliographical references (leaves 138-152).

The properties of factors present in boiled yeast juice which stimulate the oxidative breakdown of citrate in baker's yeast have been investigated. The activity has been shown to reside predominantly in the nucleotide fractions (i.e., pyridine codehydrogenases) and to a lesser extent in the ammonium salts present in whole yeast cells. The activity of the Citrate Oxidation Factor (COF) concentrated from yeast extracts and described by Foulkes (Biochem. J. 54 323 (1953)) was attributable entirely to the ammonium content of COF preparations. This activating effect of ammonium ions is accompanied by their removal from the assay system and the simultaneous production of glutamic acid. Correlation of the uptake of ammonium ion with the increased citrate metabolism indicates that the activity of the ammonium salts lies in their ability to promote the reductive amination of α-oxoglutarate. The mechanism(s) whereby this serves to stimulate the further breakdown of citrate to oxoglutarate involves the mediation of the pyridine codehydrogenases (DPN, TPN). These findings are discussed in relation to the known functioning of individual stains of the tricarboxylic acid cycle and to possible alternative pathways for the oxidation of citrate in yeast.

The tunneling fold (T-Fold) superfamily is a small superfamily of enzymes found in organisms encompassing all kingdoms of life. Seven members have been identified thus far. Despite sharing a common three-dimensional structure these enzymes perform very diverse chemistries. QueF is a bacterial NADPH-dependent oxidoreductase that catalyzes the reduction of the nitrile group of 7-cyano-7-deazaguanine (preQ0) to a primary amine (preQ1) in the queuosine biosynthetic pathway. Previous work on this enzyme has revealed the mechanism of reaction but the cofactor binding residues remain unknown. The experiments discussed herein aim to elucidate the role of residues lysine 80, lysine 83, and arginine 125 (B. subtilis numbering) in NADPH binding. The biological role of the disulfide bond between the conserved residues cysteine 55 and cysteine 99 observed in several crystal structures is also examined. Characterization of QueF mutants K80A, K83, R125A and R125K revealed lysine 80, lysine 83 and arginine 125 are required for turnover. Further analysis of turnover rates for R125K are consistent with this residue and both lysines being involved in cofactor binding presumably by interacting with the negatively charged phosphate tail of NADPH and are therefore involved in cofactor binding. Based on bond angles and energies, the disulfide bond between Cys55 and Cys99 was characterized as non-structural. Enzyme oxidation assays were consistent with the bond serving to protect QueF against irreversible oxidation of Cys55, which would render the enzyme inactive. This is the only known example of a stress protective mechanism in the Tunneling-fold superfamily. QueF-like is an amidinotransferase found in some species of Crenarchaeota and involved in the biosynthesis of archaeosine-tRNA. The work presented here is focused on the preliminary characterization of this enzyme, including the elucidation of the natural substrate as well as the source of ammonia. The structure of the enzyme was solved and is also discussed. Substrate analysis for QueF-like indicated this enzyme is capable of binding both preQ0 and preQ0-tRNA and reacting to form a thioimide intermediate analogous to QueF but only the latter serves as a substrate for the reaction. This makes QueF-like the first example of a nucleic acid binding enzyme in the Tunneling-fold superfamily. Ammonia, glutamine and asparagine were tested as nitrogen sources and unlike most known amidotransferases, QueF-like can only use free ammonia to produce the archaeosine-tRNA product. The crystal structure of P. calidifontis QueF-like indicates the functional enzyme is a dimer of pentamers pinned together by a large number of salt bridges. The structure presents a high degree of similarity to that of QueF albeit the higher twist of the QueF-like pentamers with respect to QueF results in a more compact structure.