vii, 143 leaves : ill. ; 25 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.)-- University of Adelaide, Dept. of Agricultural Biochemistry, Waite Institute, 1972

Thesis (Ph.D. -- University of Adelaide, Dept. of Agricultural Biochemistry, Waite Institute, 1972.

No description available.

No description available.

Thesis advisor: Evan R. Kantrowitz / Nucleotides, the building blocks for nucleic acids, are essential for cell growth and replication. In E. coli the enzyme responsible for the regulation of pyrimidine nucleotide biosynthesis is aspartate transcarbamoylase (ATCase), which catalyzes the committed step in this pathway. ATCase is allosterically inhibited by CTP and UTP in the presence of CTP, the end products of the pyrimidine pathway. ATP, the end product of the purine biosynthetic pathway, acts as an allosteric activator. ATCase undergoes the allosteric transition from the low-activity and low-affinity T state to the high-activity and high-affinity R state upon the binding of the substrates. In this work we were able to trap an intermediate ATCase along the path of the allosteric transition between the T and R states. Both the X-ray crystallography and small-angle X-ray scattering in solution clearly demonstrated that the mutant ATCase (K164E/E239K) exists in an intermediate quaternary structure shifted about one-third toward the canonical R structure from the T structure. The structure of this intermediate ATCase is helping to understand the mechanism of the allosteric transition on a molecular basis. In this work we also discovered that a metal ion, such as Mg2+, was required for the synergistic inhibition by UTP in the presence of CTP. Therefore, the metal ion also had significant influence on how other nucleotides effect the enzyme. A more physiological relevant model was proposed involving the metal ion. To better understand the allosteric transition of ATCase, time-resolved small-angle X-ray scattering was utilized to track the conformational changes of the quaternary structure of the enzyme upon reaction with the natural substrates, PALA and nucleotide effectors. The transition rate was increased with an increasing concentration of the natural substrates but became over one order of magnitude slower with addition of PALA. Addition of ATP to the substrates increased the rate of the transition whereas CTP or the combination of CTP and UTP exhibited the opposite effect. In this work we also studied E. coli dihydroorotase (DHOase), which catalyzes the following step of ATCase in the pyrimidine biosynthetic pathway. A virtual high throughput screening system was employed to screen for inhibitors of DHOase, which may become potential anti-proliferation and anti-malarial drug candidates. Upon the discovery of the different conformations of the 100's loop of DHOase when substrate or product bound at the active site, we've genetically incorporated an unnatural fluorescent amino acid to a site on this loop in the hope of obtaining a better understanding of the catalysis that may involve the movement of the 100's loop. / Thesis (PhD) — Boston College, 2012. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Chemistry.

Thesis (M.A.)--Boston University / In studies on the formation of 2,3-diphosphoglycerate (2,3-DPG) from inosine by erythrocyte preparations it was observed that the inosine interfered with the enzymatic assay for 2,3-DPG. Phosphoglyceromutase, for which 2,3-DPG is a coenzyme, and enolase were used in a coupled assay. The inosine appeared to interfere by inhibiting the enolase [TRUNCATED]

Thesis (Ph.D.)--Boston University / PLEASE NOTE: Boston University Libraries did not receive an Authorization To Manage form for this thesis or dissertation. It is therefore not openly accessible, though it may be available by request. If you are the author or principal advisor of this work and would like to request open access for it, please contact us at open-help@bu.edu. Thank you. / In Neurospora, as in other organisms, arginine and pyrimidine biosynthesis shares the common precursor, carbamyl phosphate. The fact that some mutants require only pyrimidines for growth while others require only arginine for growth indicates the possibility of two independent modes for carbamyl phosphate synthesis and utilization. The existence of two carbamyl phosphokinases has been hypothesized to resolve the interplay of nutritional requirements for arginine and pyrimidine. One carbamyl phosphokinase catalyzes the formation of carbamyl phosphate specific for the arginine pathway and the other carbamyl phosphokinase catalyzes the formation of carbamyl phosphate specific for the pyrimidine pathway [TRUNCATED] / 2031-01-01

No description available.

No description available.

Like its enteric counterpart, aspartate transcarbamoylase (ATCase) from Pseudomonas putida is a dodecamer of two different polypeptides. Unlike the enterics, the Pseudomonas ATCase lacks regulatory polypeptides but employs instead inactive dihydroorotases for an active dodecamer. Previous work showed that PyrB contains not only the active site but also the effector binding sites for ATP, UTP and CTP at its N-terminus. In this work, 11 amino acids were deleted from the N-terminus of PyrB and the ATCase with the truncated protein was expressed in E. coli pyrB- and purified. The wild type enzyme was similarly treated. Velocity-substrate plots without effectors gave Michaelis-Menten kinetics in all cases. Deleting 11 amino acids did not affect dodecameric assembly but altered effector responses. When carbamoylphosphate was varied, the mutant enzyme was inhibited by UTP while the wild type enzyme was activated 2-fold. When the aspartate was varied, CTP had no effect on the mutant enzyme but strongly inhibited the wild type enzyme.