The thermoalkaliphilic Bacillus sp. strain TA2.A1 was able to grow in pH-controlled batch culture containing a fermentable growth substrate (i.e. sucrose) from pH 7.5 to 10.0 with no significant change in specific growth rate, suggesting that this bacterium is a facultative alkaliphile. However, when strain TA2.A1 was grown on non-fermentable carbon sources like succinate, no growth was observed until the external pH was > 9.0, suggesting this bacterium is an obligate alkaliphile. Growth on succinate at pH 9.5 was sensitive to both carbonyl cyanide m-chlorophenylhydrazone (CCCP) and monensin revealing that both the proton and sodium motive force ([Delta][mu][H⁺] and [Delta][mu][Na⁺], respectively) were obligate requirements for growth at alkaline pH values. Transport of succinate was driven by a chemical gradient of Na⁺ ([Delta]pNa⁺) that was strictly coupled to [Delta][Psi]. A single transport system was detected for the uptake of succinate, with an apparent K[m] of 19 [mu]M and V[max] of 0.45 nmol succinate/min/mg protein. Succinate transport was pH-dependent, and showed optimal activity at pH values greater than 8.5. Other C₄-dicarboxylates (e.g. malate, fumarate) inhibited the uptake of succinate suggesting that the permease was general for other C₄-dicarboxylates. Cytochrome content, succinate dehydrogenase oxidoreductase, and F₁F₀-ATPase activities were lower in membranes from strain TA2.A1 cells grown at pH 7.5 compared to those cultured at 9.5. These data suggest that oxidative phosphorylation-linked processes are down-regulated at neutral pH values, an observation that mirrored oxygen consumption profiles of strain TA2.A1 in whole cells. To study this phenomenon at a molecular level, we measured ATP synthesis by the F₁F₀-ATP synthase from strain TA2.A1 as a function of pH. The strain TA2.A1 F₁F₀-ATP synthase had a pH optimum for ATP synthesis of 9.0-9.5, and significantly lower rates of ATP synthesis observed below pH 9.0. Analysis of the atp operon from the thermoalkaliphilic Bacillus sp. strain TA2.A1 and comparison with other atp operons from alkaliphilic bacteria reveals the presence of a conserved lysine residue at position 180 (Bacillus sp. strain TA2.A1 numbering) within the a subunit of these F₁F₀-ATP synthases. We hypothesize that the basic nature of this residue is ideally suited to capture protons from the bulk phase at high pH. To test this hypothesis, a heterologous expression system for the ATP synthase from Bacillus sp. TA2.A1 (TA2F₁F₀) was developed in Escherichia coli DK8 ([Delta]atp). Amino acid substitutions were made in the a subunit of TA2F₁F₀ at position 180. Lysine (aK180) was substituted for the basic residues histidine (aK180H) or arginine (aK180R), and the uncharged residue glycine (aK180G). ATP synthesis experiments were performed in ADP plus P[i]-loaded right-side out membrane vesicles energized by ascorbate-phenazine methosulfate. When these enzyme complexes were examined for their ability to perform ATP synthesis over the pH range from 7.0 to 10.0, TA2F₁F₀ and aK180R showed a similar pH profile having optimum ATP synthesis rates at pH 9.0 to 9.5 with no measurable ATP synthesis at pH 7.5. Conversely, aK180H and aK180G showed maximal ATP synthesis at pH�s 8.0 and 7.5, respectively. ATP synthesis under these conditions for all enzyme forms was sensitive to DCCD. These data strongly imply that amino acid residue K180 is a specific adaptation within the a subunit of TA2F₁F₀ to facilitate proton capture at high pH. At pH values near the pK[a] of K180, the trapped protons readily dissociate to reach the subunit c binding sites but this dissociation is impeded at neutral pH values causing either a blocking of the proposed H⁺ channel and/or mechanism of proton translocation, and hence ATP synthesis is inhibited. The mechanisms where by alkaliphilic bacteria obtain iron remains unknown. Growth of strain TA2.A1 at pH 9.5 in the presence of the artificial iron chelators ethylenediamine O-hydroxyphenylacetic acid (EDDHA) and 2�2� dipyridal revealed that iron is an important requirement for aerobic growth at alkaline pH values. Furthermore, biochemical analysis showed that Bacillus alcalophilus and Bacillus pseudofirmus both synthesized orange catecholate siderophores, whilst Bacillus halodurans synthesized a hydroxamate siderophore. These tests showed that strain TA2.A1 synthesized both orange catecholate and hydroxamate siderophore/s. Attempts to purify the catecholate were unsuccessful. No homologues of previously identified non-ribosomal peptide synthase (NRPS) genes in Bacillus subtilis and B.halodurans were detected in the genome of strain TA2.A1 using both PCR and Southern hybridization using known non-ribosomal peptide synthase genes.

Two identical instruments have been developed for use in the field to make radiometric measurements. They have been described as spectropolarimeters because of their ability to make polarization measurements in narrow spectral passbands. They have been used as part of a NASA sponsored project to monitor the spectral and temporal response of the thematic mapper satellites. These satellites allow many natural and man-tended resources to be monitored over years of time, thus allowing their use to be planned for in the future. The dissertation discusses the design, fabrication, testing and absolute radiometric calibration of these spectropolarimeter instruments. The outstanding feature of these instruments are that they have been calibrated absolutely, for radiance measurements, to an accuracy of 2% - 3%, in the range of 400 nm to 1040 nm over selected spectral passbands. Previously, field measurements were considered good if they had an absolute accuracy of 10%, implying that the present accuracies are advancing the state-of-the-art for field instrument calibrations. These improved accuracies are the result of using two recently developed calibration standards, both of which use detector based methods. These standards are the Electrically Calibrated Pyroelectric Radiometer (ECPR), and the QED-100 quad detector. The end of the dissertation discusses the attempts made to verify that the accuracies claimed are indeed valid, and it is the author's belief that these accuracies have been verified completely.

published_or_final_version / Physiology / Doctoral / Doctor of Philosophy

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No description available.

This thesis describes an investigation of how techniques of modelling, estimation and control can be used to improve regulation of pH. A novel mathematical model of neutralization is used which explicitly accounts for two separate sources of non-linearities in pH systems: the inherent non-linearity of the pH measurement and the variable and uncertain non-linearity of the neutralization chemistry. A detailed interactive simulation package was developed. It has been used to study: <ul><li>1. The application of a Bayes' non-linear estimator to a noisy pH measurement. The estimator is used in conjunction with an Extended Kalman Filter in order to generate estimates for use in combined feedforward/feedback controllers. A novel version of the Bayes' estimator is developed which successfully estimated jumping and drifting bias on the pH measurement.</li><li>2. A novel cautious stochastic control law which uses the generated estimates and allows for their uncertainty. Reduced quality of estimation forces the controller to regulate the pH at an optimally tuned target value where control is easier.</li><li>3. Comparisons of a variety of modern and conventional control algorithms. The modern algorithms include three self-tuners, the non-linear estimators and the cautious controller. The conventional algorithms include a PID controller, analytical linearisation of the pH measurement and the use of estimated or measured feedforward signals. Twenty-five pH control laws are simulated.</li></ul> The quality of regulation which can be achieved depends on what prior information is available about the neutralisation process. A strategy is proposed which matches controller structure to available information. This strategy provides guidelines to the control configurations that can be implemented as well as their relative limitations. It is shown that improvement in performance over a PID regulator is possible if one uses more complex controllers. This improvement is investigated by simulation because it varies from one process to another. It is also shown how the resulting simulation package has been used for computer aided design of pH control systems in the chemical industry.

The thermoalkaliphilic Bacillus sp. strain TA2.A1 was able to grow in pH-controlled batch culture containing a fermentable growth substrate (i.e. sucrose) from pH 7.5 to 10.0 with no significant change in specific growth rate, suggesting that this bacterium is a facultative alkaliphile. However, when strain TA2.A1 was grown on non-fermentable carbon sources like succinate, no growth was observed until the external pH was > 9.0, suggesting this bacterium is an obligate alkaliphile. Growth on succinate at pH 9.5 was sensitive to both carbonyl cyanide m-chlorophenylhydrazone (CCCP) and monensin revealing that both the proton and sodium motive force ([Delta][mu][H⁺] and [Delta][mu][Na⁺], respectively) were obligate requirements for growth at alkaline pH values. Transport of succinate was driven by a chemical gradient of Na⁺ ([Delta]pNa⁺) that was strictly coupled to [Delta][Psi]. A single transport system was detected for the uptake of succinate, with an apparent K[m] of 19 [mu]M and V[max] of 0.45 nmol succinate/min/mg protein. Succinate transport was pH-dependent, and showed optimal activity at pH values greater than 8.5. Other C₄-dicarboxylates (e.g. malate, fumarate) inhibited the uptake of succinate suggesting that the permease was general for other C₄-dicarboxylates. Cytochrome content, succinate dehydrogenase oxidoreductase, and F₁F₀-ATPase activities were lower in membranes from strain TA2.A1 cells grown at pH 7.5 compared to those cultured at 9.5. These data suggest that oxidative phosphorylation-linked processes are down-regulated at neutral pH values, an observation that mirrored oxygen consumption profiles of strain TA2.A1 in whole cells. To study this phenomenon at a molecular level, we measured ATP synthesis by the F₁F₀-ATP synthase from strain TA2.A1 as a function of pH. The strain TA2.A1 F₁F₀-ATP synthase had a pH optimum for ATP synthesis of 9.0-9.5, and significantly lower rates of ATP synthesis observed below pH 9.0. Analysis of the atp operon from the thermoalkaliphilic Bacillus sp. strain TA2.A1 and comparison with other atp operons from alkaliphilic bacteria reveals the presence of a conserved lysine residue at position 180 (Bacillus sp. strain TA2.A1 numbering) within the a subunit of these F₁F₀-ATP synthases. We hypothesize that the basic nature of this residue is ideally suited to capture protons from the bulk phase at high pH. To test this hypothesis, a heterologous expression system for the ATP synthase from Bacillus sp. TA2.A1 (TA2F₁F₀) was developed in Escherichia coli DK8 ([Delta]atp). Amino acid substitutions were made in the a subunit of TA2F₁F₀ at position 180. Lysine (aK180) was substituted for the basic residues histidine (aK180H) or arginine (aK180R), and the uncharged residue glycine (aK180G). ATP synthesis experiments were performed in ADP plus P[i]-loaded right-side out membrane vesicles energized by ascorbate-phenazine methosulfate. When these enzyme complexes were examined for their ability to perform ATP synthesis over the pH range from 7.0 to 10.0, TA2F₁F₀ and aK180R showed a similar pH profile having optimum ATP synthesis rates at pH 9.0 to 9.5 with no measurable ATP synthesis at pH 7.5. Conversely, aK180H and aK180G showed maximal ATP synthesis at pH�s 8.0 and 7.5, respectively. ATP synthesis under these conditions for all enzyme forms was sensitive to DCCD. These data strongly imply that amino acid residue K180 is a specific adaptation within the a subunit of TA2F₁F₀ to facilitate proton capture at high pH. At pH values near the pK[a] of K180, the trapped protons readily dissociate to reach the subunit c binding sites but this dissociation is impeded at neutral pH values causing either a blocking of the proposed H⁺ channel and/or mechanism of proton translocation, and hence ATP synthesis is inhibited. The mechanisms where by alkaliphilic bacteria obtain iron remains unknown. Growth of strain TA2.A1 at pH 9.5 in the presence of the artificial iron chelators ethylenediamine O-hydroxyphenylacetic acid (EDDHA) and 2�2� dipyridal revealed that iron is an important requirement for aerobic growth at alkaline pH values. Furthermore, biochemical analysis showed that Bacillus alcalophilus and Bacillus pseudofirmus both synthesized orange catecholate siderophores, whilst Bacillus halodurans synthesized a hydroxamate siderophore. These tests showed that strain TA2.A1 synthesized both orange catecholate and hydroxamate siderophore/s. Attempts to purify the catecholate were unsuccessful. No homologues of previously identified non-ribosomal peptide synthase (NRPS) genes in Bacillus subtilis and B.halodurans were detected in the genome of strain TA2.A1 using both PCR and Southern hybridization using known non-ribosomal peptide synthase genes.

Thesis (M.S.)--University of Missouri-Columbia, 2007. / The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Title from title screen of research.pdf file (viewed on April 3, 2009) Includes bibliographical references.

V.G. Behrmann's thesis (PH. D.)--University of Michigan, 1939. / Cover-title. "Reprinted from the American journal of physiology, vol. 132, no. 1, February, 1941." eContent provider-neutral record in process. Description based on print version record. "References": p. 280.

Thesis (Ph. D.)--University of Virginia, 1928.