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 Title: Experimental and theoretical studies of the peroxidase-NADH biochemical oscillator: An enzyme-mediated chemical switch Author(s): Olson, Dean Lee Doctoral Committee Chair(s): Scheeline, Alexander Department / Program: Chemistry Discipline: Chemistry Degree Granting Institution: University of Illinois at Urbana-Champaign Degree: Ph.D. Genre: Dissertation Subject(s): Chemistry, Analytical Chemistry, Biochemistry Chemistry, Physical Abstract: The peroxidase-NADH system is one of the few biochemical examples among known in vitro chemical oscillators. This system functions as a model for the development of methods and principles to study the nonlinear dynamics of cyclic, biochemical reactions. Experiments are conducted on a standard oscillator, which consists of a well-mixed, aqueous solution of the enzyme horseradish peroxidase and the common electron transfer dye, methylene blue (MB$\sp+),$ to which $\beta$-nicotinamide adenine dinucleotide (NADH) and oxygen are added at constant rates. The concentrations of several chemical species show prolonged oscillations over time.A multidimensional approach to data acquisition is used to provide an improved experimental basis for more realistic modeling. Oscillatory dynamics were investigated, including oxygen mass transport, the role of MB$\sp+,$ intermediates hydrogen peroxide $\rm(H\sb2O\sb2)$ and superoxide (O$\sb2\sp{\bullet-}),$ damping, and the effect of light used for absorbance measurements. MB$\rm\sp+, H\sb2O\sb2,$ and O$\sb2\sp{\bullet-}$ are essential for oscillations in both experimental and theoretical studies. MB$\sp+$ is shown in experiments to inhibit the consumption of oxygen.To simulate experimental results, a minimal model is developed which incorporates chemically realistic parameters. The model consists of thirteen reactions and rate constants, and fifteen species. Nine of the rate constants are from the published literature, and four originate from experimental measurements described here. In the model, two reactions of superoxide perform a key function as an enzyme-mediated chemical switch. In oscillatory simulations, the oxidation of Per$\sp{3+}$ by superoxide is the controlling reaction when the oxygen level increases (the rising portion of an oscillation). When the Per$\sp{3+}$ is depleted and has been transformed to Per$\sp{6+},$ the reaction switches to superoxide disproportionation, resulting in a net decrease in oxygen. Once the Per$\sp{6+}$ has been converted back to the native Per$\sp{3+}$ by supporting reactions, the system switches again to Per$\sp{3+}$ oxidation by superoxide, and the cycle restarts.The success of the straightforward approach used to build the model demonstrates that chemical knowledge can be applied to create an oscillatory system without invoking abstract models as a starting point. The postulated kinetic scheme presented here has its greatest potential as a chemically realistic bridge between experiment and theory. Issue Date: 1994 Type: Text Language: English URI: http://hdl.handle.net/2142/21044 Rights Information: Copyright 1994 Olson, Dean Lee Date Available in IDEALS: 2011-05-07 Identifier in Online Catalog: AAI9512502 OCLC Identifier: (UMI)AAI9512502
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