Thesis (Ph. D.)--Washington State University, May 2009. / Title from PDF title page (viewed on July 31, 2009). "School of Biological Sciences." Includes bibliographical references.

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

Thesis (Ph. D.)--University of Wisconsin--Madison, 1984. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographies.

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

Thesis (Ph. D.)--University of Oregon, 2005. / Typescript. Includes vita and abstract. Includes bibliographical references (leaves 85-95). Also available for download via the World Wide Web; free to University of Oregon users.

ATP synthase (F_1 F_O-ATPase) is an essential enzyme for life. Powered by an electrochemical proton gradient, it catalyzes ADP and phosphate into ATP. The F_1-subunit of ATP synthase is called F_1-ATPase as it also independently catalyzes the reverse reaction in absence of F_O-part. The nearly 100% energy conversion efficiency of the molecular motor has attracted the attention of many physicists and biologists to explore the underlying thermodynamics. Recently, a new nonequilibrium equality derived by Harada and Sasa (Harada & Sasa, 2005) was applied to the experimental time series data on F_1-ATPase to extract heat flow to the environment. A phenomenological model for rotary motion was proposed and shown to reproduce key experimental features. Interested in the high efficiency of F_1-ATPase and the good performance of the corresponding model, we carried out a detailed computational study of the model to understand its behavior in a broader range of parameter values. We solved the model using a modified Gillespie algorithm for stochastic simulation and by integrating the Fokker-Planck equation. Various physical properties of the model, such as the relation between rotational velocity and parameters characterizing angular dependence (q) and ATP switching rates (W), the relation between two kinds of dissipation and rotational velocity, the negative heat flow from environment to system through ATP binding etc. are analyzed in detail. Importantly, we modified the driving potential to investigate the factors affecting the efficiency. Additionally, we found some inconsistences between properties of this model and previous studies and we could unify them by some adjustments, which may be useful for constructing more precise models in the future.

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Some of the kinetic characteristics of the sodium and potassium-activated adenosine triphosphatase of the fragmented cell membranes of cells from the gills of sea water adapted coho salmon Oncorhynchus kisutch, and changes in this enzyme upon exposure to sea water and during the fry to smolt stages of fresh water reared juvenile coho were investigated. Inhibition with 4 X 10⁻⁴ moles/liter ouabain was used to assay the activity of this enzyme since this ATPase is specifically inhibited by ouabain (Skou, 1957). The following assay conditions were found to result in maximal hydrolysis of ATP in enzyme preparations from sea water adapted coho: pH, 7.4; incubation temperature, 40°C; NaCl and KCl concentrations of 100.0 and 20.0 mmoles/liter, respectively, and Mg²⁺ -ATP, 5.0 mmoles/liter. The Km for ATP was 0.2 mmoles/liter. The enzyme activity recorded with magnesium ions as the only cation present (Mg²⁺-ATPase) was not affected by any concentration of ouabain, although the addition of sodium ions (100 mmoles/liter) appeared to inhibit this activity slightly. The additional hydrolysis of ATP observed when sodium, potassium and magnesium ions were present was inhibited by ouabain. The Ki for ouabain was; 7 X 10⁻⁶ moles/liter when sodium and potassium ion concentrations were 100.0 and 20.0 mmoles liter, respectively. The (Na⁺ + K⁺)- activated ATPase of sea water adapted coho was characterized by its high ouabain-sensitive activity and the large activating effect of potassium ions in the presence of magnesium and sodium ions compared to the activity observed with the latter two ions alone. This enzyme in preparations from the gills of fresh water reared fish was characterized by a high activating effect of sodium ions when present with magnesium ions. This sodium activation often comprised over 60% of the total ouabain-sensitive activity. Considerable increases in the total activity, and activating effects of potassium ions and decreases in the activating effects of sodium ions alone were observed when fresh water reared coho were transferred directly to sea water. The changes in the activating effect of the ions were noticable after 5 days exposure to sea water although no changes in the total activity of the enzyme occurred until after 10 days exposure. On a seasonal basis changes in enzyme activity occurred which were apparently linked to the stage of development of the parr-smolt transformation in fresh water reared juvenile coho. Activities during the period of October 1, 1968 to late November, 1968 were generally quite low. A sharp peak in activity occurred in December, 1968 to late January, 1969 which decreased to a low level by mid-February. Up to and including this last period the activity of enzymes from the gills of both fresh water and sea water reared coho were qualitatively similar although the seawater fish always had a higher enzyme activity. During the period of mid-February to late April,1969 the enzyme from fresh water reared coho changed in total activity and characteristics of sodium activation and potassium activation and became very similar to that of sea water reared fish of the same age. / Science, Faculty of / Zoology, Department of / Graduate

The ATPase complex of E. coll consists of two functional units. ECF[sub=1] is an extrinsic membrane protein having the active site(s) for ATP synthesis and hydrolysis. F₀ is intrinsic and catalyzes the reversible transfer of protons across the membrane. ECF[sub=1] consists of five polypeptides (α- ε) ranging in molecular weight from 13 000 - 57 000. F₀ has three polypeptides (9 000, 18 000, 24 000), the smallest of which is the dicyclohexylcarbodiimide (DCCD)-binding protein postulated to be a transmembrane pathway for proton translocation. An ECF₁F₀ complex was solubilized from the membranes of E. coli with N-lauroyl sarcosine and purified by chromatography on Phenyl-Sepharose CL-4B followed by sedimentation of the enzyme at 250 000 xg for 16-17 h. The purified ECF₁F₀ complex consisted of the eight polypeptides described above, as well as associated polypeptides of molecular weights 30 000, 28 000 and 14 000. Removal of ECF₁ from the membranes of the wild-type E. coli resulted in the membranes becoming leaky to protons so that they could not be energized. The unc mutants, E. coli AN382, CBT-302 and N₁₄₄ could maintain a proton gradient across the membrane in the absence of ECF₁. A normal DCCD-binding protein was present in the F₀ complex of each mutant. However, the 18 000 dalton polypeptide of F₀ was absent in the membranes of E. coli N₁₄₄, suggesting that it was required for a functional F₀. The involvement of the 18 000 dalton polypeptide in the proton-translocating activity was also suggested by the observation that this polypeptide was absent in the ECF₁F₀ complex immunoprecipitated from trypsin-treated "stripped" vesicles, which had been reconstituted with ECF₁. Although these trypsin-treated "stripped" vesicles could rebind ECF₁, the membranes could not be energized during ATP hydrolysis. Leakiness of the membranes to protons could be repaired by the reaction of the ECF₁ stripped membranes with DCCD or ECF₁. Similarly, antibody raised against the DCCD-binding protein prevented this leakage of protons. The antibody also inhibited the rebinding of ECF₁ to the "stripped" everted membrane vesicles. These results indicated that the DCCD-binding protein was exposed on the cytoplasmic surface of the cell. Attempts to show whether the DCCD-binding protein was transmembranous were not successful. Radioimmunoassay techniques were used to show In vitro, the involvement of the arginyl residue(s) of the DCCD-binding protein in the binding of ECF₁. Binding of ECF₁ to the DCCD-binding protein appeared to involve the α and/or β subunits of ECF₁. Chemical modification of the methionyl residue(s) of the DCCD-binding protein did not alter its capacity to bind ECF₁, but destroyed the antigenic site(s) of the polypeptide. In summary, these results are consistent with the proposed "loop" arrangement of the DCCD-binding protein in which the polar central region of this molecule is at the cytoplasmic surface of the cell membrane. / Medicine, Faculty of / Biochemistry and Molecular Biology, Department of / Graduate