No description available.

Dissertation (Ph.D.)--Auburn University, 2007. / Abstract. Includes bibliographic references (p.147-156).

Thesis (Ph. D.)--Ohio State University, 2004. / Document formatted into pages. Includes bibliographical references. Abstract available online via OhioLINK's ETD Center; full text release delayed at author's request until 2005 Aug. 18.

Thesis (Ph. D.)--Ohio State University, 2003. / Title from first page of PDF file. Document formatted into pages; contains xxi, 250 p.; also includes graphics (some col.) Includes bibliographical references (p. 226-250). Available online via OhioLINK's ETD Center

Thesis (Ph. D.)--Ohio State University, 2007. / Title from first page of PDF file. Includes bibliographical references (p. 181-195).

Thesis (Ph.D.)--University of Delaware, 2009. / Principal faculty advisor: George W. Luther, III., College of Earth, Ocean, & Environment. Includes bibliographical references.

Iron sulfur [Fe-S] proteins are essential components of many major biological processes including electron transport, respiration, photosynthesis, hormone biosynthesis, and environmental sensing. The process of [Fe-S] cluster assembly in living cells is a controlled mechanism that is highly conserved across all kingdoms. Considerable progress has been made in deciphering this mechanism in bacteria, yeast, and mammals. The key players are the NifS/IscS/SufS proteins, which act as the sulfur donor, and the NifU/IscU/SufU proteins, which serve as a scaffold that binds Fe and upon which the cluster is assembled. Additional proteins are involved in the maturation and transport of the clusters. In eukaryotes there is redundancy in the proteins involved in this mechanism and the process is compartmentalized. Not much is known about the [Fe-S] cluster assembly mechanism in plants. In addition to the redundancy and compartmentalization seen in this machinery in eukaryotes, plants present a further challenge by offering chloroplasts as an additional site for [Fe-S] cluster assembly. The objective of this project has been to characterize Arabidopsis AtNFS1 and AtISU1-3, which show high homology to NifS/IscS and NifU/IscU, respectively, and are hypothesized to be key players in [Fe-S] cluster biogenesis in plants. Subcellular localization results of the AtNFS1 and AtISU1-3 proteins fused to GFP from this study are consistent with the presence of dual machinery in plants, with both mitochondria and chloroplasts as sites for [Fe-S] cluster assembly. Furthermore, observations also showed that AtISU2 mRNA may be unstable. The results of these experiments, together with promoter analysis described in this dissertation using GUS fusions suggested that the genes encoding the AtISU scaffold proteins are regulated at the transcriptional and probably also at the posttranscriptional level. Gene silencing experiments performed in this dissertation research using antisense and RNAi constructs indicated that these genes have the potential to impact respiration, photosynthesis, phytohormone biosynthesis, and environmental sensing, diverse processes that rely on [Fe-S] proteins. These observations, together with previous in vitro evidence that AtNFS1 and AtISU1 can participate in [Fe-S] cluster assembly, provide strong evidence that these proteins are part of two distinct cluster assembly systems that function in different subcellular locations and perhaps under different environmental conditions. Information gathered here has made it possible to begin developing a detailed model of [Fe-S] cluster biogenesis in plants. / Ph. D.

Inaug.-Diss.--Giessen. / Vita.

The iron-sulfur flavoprotein (Isf) from the acetate utilizing methanoarchaeon Methanosarcina thermophila was heterologously produced in Escherichia coli, purified to homogeneity, and characterized to determine the properties of the iron-sulfur cluster and FMN. Chemical and spectroscopic analyses indicated that Isf contained one 4Fe-4S cluster and one FMN per monomer. The midpoint potentials of the [4Fe-4S]2+/1+ center and FMN/FMNH2 redox couple were -394 and -277 mV respectively. The deduced amino acid sequence of Isf revealed high identity with Isf homologues from the CO2 reducing methanoarchaea Methanococcus jannaschii and Methanobacterium thermoautotrophicum. Extracts of H2-CO2-grown M. thermoautotrophicum cells were able to reduce Isf from M. thermophila using either H2 or CO as the reductant. Addition of ferredoxin A to the reaction further stimulated the rate of Isf reduction. These results suggest that Isf homologues are coupled to ferredoxin in electron transfer chains in methanoarchaea with diverse metabolic pathways. Reconstituted systems containing carbon monoxide dehydrogenase/acetyl-CoA synthase complex (CODH/ACS), ferredoxin A, Isf, and the designated electron carriers (NAD, NADP, F420, and 2-hydroxyphenazine) were used in an attempt to determine the electron acceptor for Isf. Isf was unable to reduce any of these compounds. Furthermore, 2-hydroxyphenazine competed with Isf to accept electrons from ferredoxin A indicating that ferredoxin A is a more favorable electron partner for 2-hydroxyphenazine. Thus, the physiological electron acceptor for Isf is unknown. Amino acid sequence alignment of Isf sequences revealed a conserved atypical cysteine motif with the potential to ligate the 4Fe-4S cluster. Site-directed mutagenesis of the cysteine residues in this motif, and the two additional cysteines in the sequence, was used to investigate these cysteine residue as ligands for coordinating the 4Fe-4S center of Isf. Spectroscopic and biochemical analyses were consistent with the conserved cysteine motif functioning as ligating the 4Fe-4S center. Redox properties of the 4Fe-4S and FMN centers revealed a role for the 4Fe-4S center in the transfer of electrons from ferredoxin A to FMN. / Ph. D.

No description available.