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dc.contributor.advisorDuin, Evert
dc.contributor.authorEngel, Carly
dc.date.accessioned2019-07-17T16:42:38Z
dc.date.available2019-07-17T16:42:38Z
dc.date.issued2019-07-17
dc.identifier.urihttp://hdl.handle.net/10415/6835
dc.description.abstractGlobal warming is a continuing crisis today. It has caused great damage to our planet and health. One of the leading causes of this issue originates from humans releasing an enormous amount of greenhouse gases into the air, in particular CO2 and CH4. Infrared radiation produced by the sun become trapped by these molecules and, therefore, increases the Earth’s temperature drastically. Ultimately, the source of most of the CH4 is biological. One enzyme known to aid in the production of methane is heterodisulfide reductase (Hdr). Hdr recycles the substrates used by the enzyme that catalyzes the methane formation. In addition, reduced ferredoxin (FdRED) is produced and used to push the first step in methanogenesis. The goal of this research is to understand the role of the cofactors, iron-sulfur clusters, and flavin adenine dinucleotide (FAD) in catalysis, with emphasis on the flavin-based electron bifurcation process and heterodisulfide (HDS) reduction. Redox titrations of methyl-viologen hydrogenase: heterodisulfide reductase (Mvh:Hdr) yielded multiple midpoint potentials for different [4Fe-4S]+ and [2Fe-2S]+ clusters. In addition, rapid freeze quench was performed on the whole complex. This allowed the detection of several redox active species, calculation of midpoint potential (Em), and the development of a model of Hdr with assignment of electron paramagnetic resonance (EPR) signal and Em values to specific cofactors in the Hdr complex. Due to the presence of a total of 14 clusters it was not always possible to discern individual signals in the whole Mvh:Hdr complex. Therefore, similar experiments were executed on the isolated HdrB, and HdrBC subunits. The HdrB subunit is the site of the HDS reduction. HdrB2 and HdrB2C2 were cloned by Dr. James “Greg” Ferry’s group from Methanosarcina acetivorans into Escherchia coli. Two non-cuboidal clusters were expected in the HdrB subunit. Our preparation contained one [2Fe-2S] cluster and one non-cuboidal [4Fe-4S] cluster. Through further site-directed mutagenesis both sites could be studied separately. The data showed that the non-cuboidal [4Fe-4S] clusters do not show a 1+/2+ transition. In the presence of substrate, a 2+/3+ transition is detected. Surprisingly the HdrB2 and HdrB2C2 preparations are active although only one non-cuboidal cluster is present.en_US
dc.rightsEMBARGO_NOT_AUBURNen_US
dc.subjectChemistry and Biochemistryen_US
dc.titleProperties of Iron-Sulfur Clusters in Heterodisulfide Reductaseen_US
dc.typePhD Dissertationen_US
dc.embargo.lengthMONTHS_WITHHELD:25en_US
dc.embargo.statusEMBARGOEDen_US
dc.embargo.enddate2021-07-30en_US
dc.contributor.committeeGoodwin, Doug
dc.contributor.committeeEllis, Holly
dc.contributor.committeeGorden, Anne
dc.contributor.committeePantazes, Robert


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