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Effects Of Iron Limitation And Carbon Source On Extracellular Excretions And Intracellular Carbon Metabolism In Pseudomonas Putida

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Abstract

Iron (Fe) is an important micronutrient in soils, essential for bacterial growth and carbon metabolism, serving as an enzymatic cofactor for many central carbon metabolic reactions. It has limited bioavailability due to being embedded in Fe oxide and hydroxide minerals. Bacteria secrete high-affinity metal-binding molecules, or siderophores, along with low-affinity organic acids to cope with Fe limitation and facilitate mineral dissolution. In the research presented, I investigated the role of available carbon sources (glucose, succinate, acetate, and citrate) on the secretion of these molecules from the ubiquitous soil bacteria Pseudomonas putida and their effectiveness in dissolving Fe from common soil minerals. Fe limited cells secreted the siderophore pyoverdine in addition to secreting higher levels of small organic acids. Dissolution experiments were carried out with bacterial secretions, substrates, and detected organic acids to determine the amount of Fe released with each organic compound. Overall, bacterial secretions were more effective in dissolving Fe than the individual substrates and organic acids. Additionally, to elucidate the changes in metabolic flux with respect to Fe, kinetic and steadystate isotopic labeling experiments were performed with [U-13C] and [1,2-13C] glucose under Felimited and replete conditions. Steady-state experiments were also performed with [2,3-13C] succinate under Fe-replete conditions to compare changes in metabolic flux between carbon sources. Overall, most fluxes decreased under Fe-limited conditions with glucose as the sole carbon source, but the flux to the amino acid precursor phosphoenolpyruvate increased. The succinate flux model showed a decrease in flux through the citric acid cycle under Fe-replete conditions, with a corresponding decrease in the amount of CO2 produced compared to Fereplete glucose. These findings indicate that the secretion of organic molecules is substrate and Fe dependent, as are the carbon fluxes through the pathways of the central carbon metabolism.

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2015-01-26

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Pseudomonas putida; Metabolism; Iron

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Aristilde, Ludmilla

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Walker, Larry P

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Agricultural and Biological Engineering

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M.S., Agricultural and Biological Engineering

Degree Level

Master of Science

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Government Document

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dissertation or thesis

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