By definition, aerobic organisms, both prokaryotic and eukaryotic, require molecular oxygen for energy conservation. This is a mixed blessing, however, as extremely reactive oxygen derivatives are produced during normal metabolism that can damage all cellular components. These so-called reactive oxygen species (ROS) have been implicated in a wide variety of chronic and infectious human diseases, including cancer, Alzheimer's disease, arthritis and AIDS, yet ROS are also used as a defense system against pathogens and in signal transduction pathways. Understanding the responses of microbes to oxygen has direct ramifications for the treatment of diseases caused by anaerobic pathogens. In 1999 we proposed that anaerobes have a novel response to ROS in which a non-heme iron protein termed superoxide reductase (SOR) played a key role. SOR was characterized from the hyperthermophilic anaerobe, Pyrococcus furiosus, and over the prior funding period it has been established using structural and spectroscopic approaches that SOR is uniquely suited to catalyze superoxide reduction. Using DNA microarrays to all 2065 ORFs in the complete P. furiosus genome, it was shown that the genes encoding SOR and related proteins are all expressed at significant levels in the absence of any oxidative shock. P. furiosus is therefore continuously 'armed' and ready to deal with ROS exposure. This is a first line of defense, however, as DNA microarray analyses show that the complete response to oxidative stress requires the induction of a large number of novel proteins (encoded by conserved/hypothetical genes), some of which are also induced by growth at sub-optimal temperatures. In the proposed research, the novel stress-regulated proteins, together with SOR and related reductases and oxidases, will be characterized with respect to their regulation, multiprotein complex formation, and catalytic functions using immunological, biochemical and structural analyses. A variety of complementary spectroscopic techniques, including EPR, ENDOR, MCD, resonance Raman, FTIR and X-ray absorption, will be utilized to probe the catalytic function of specific members of the stress-related pathways, with particular emphasis on SOR. The results will provide completely new insights into the stress responses of anaerobes, and provide strategies for determining the function of uncharacterized hypothetic algenes that typically account for half of a microbial genome.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM060329-04A1
Application #
6819372
Study Section
Physical Biochemistry Study Section (PB)
Program Officer
Preusch, Peter C
Project Start
2000-03-01
Project End
2008-02-28
Budget Start
2004-07-05
Budget End
2005-02-28
Support Year
4
Fiscal Year
2004
Total Cost
$221,110
Indirect Cost
Name
University of Georgia
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
004315578
City
Athens
State
GA
Country
United States
Zip Code
30602
Strand, Kari R; Sun, Chengjun; Li, Ting et al. (2010) Oxidative stress protection and the repair response to hydrogen peroxide in the hyperthermophilic archaeon Pyrococcus furiosus and in related species. Arch Microbiol 192:447-59
Hamilton-Brehm, Scott D; Schut, Gerrit J; Adams, Michael W W (2009) Antimicrobial activity of the iron-sulfur nitroso compound Roussin's black salt [Fe4S3(NO)7] on the hyperthermophilic archaeon Pyrococcus furiosus. Appl Environ Microbiol 75:1820-5
Dey, Abhishek; Jenney Jr, Francis E; Adams, Michael W W et al. (2007) Sulfur K-edge X-ray absorption spectroscopy and density functional theory calculations on superoxide reductase: role of the axial thiolate in reactivity. J Am Chem Soc 129:12418-31
Kelley, Laura-Lee Clancy; Dillard, Bret D; Tempel, Wolfram et al. (2007) Structure of the hypothetical protein PF0899 from Pyrococcus furiosus at 1.85 A resolution. Acta Crystallogr Sect F Struct Biol Cryst Commun 63:549-52
Gerwe, Brian; Kelley, Laura-Lee Clancy; Dillard, Bret D et al. (2007) Structural and transcriptional analyses of a purine nucleotide-binding protein from Pyrococcus furiosus: a component of a novel, membrane-bound multiprotein complex unique to this hyperthermophilic archaeon. J Struct Funct Genomics 8:1-10
Dey, Abhishek; Jenney Jr, Francis E; Adams, Michael W W et al. (2007) Solvent tuning of electrochemical potentials in the active sites of HiPIP versus ferredoxin. Science 318:1464-8
Cacciapuoti, Giovanna; Porcelli, Marina; Moretti, Maria Angela et al. (2007) The first agmatine/cadaverine aminopropyl transferase: biochemical and structural characterization of an enzyme involved in polyamine biosynthesis in the hyperthermophilic archaeon Pyrococcus furiosus. J Bacteriol 189:6057-67
Tan, Ming-Liang; Bizzarri, Anna Rita; Xiao, Yuming et al. (2007) Observation of terahertz vibrations in Pyrococcus furiosus rubredoxin via impulsive coherent vibrational spectroscopy and nuclear resonance vibrational spectroscopy--interpretation by molecular mechanics. J Inorg Biochem 101:375-84
Tran, Thao T; Dam, Phuongan; Su, Zhengchang et al. (2007) Operon prediction in Pyrococcus furiosus. Nucleic Acids Res 35:11-20
Wang, Xu; Lee, Han-Seung; Sugar, Frank J et al. (2007) PF0610, a novel winged helix-turn-helix variant possessing a rubredoxin-like Zn ribbon motif from the hyperthermophilic archaeon, Pyrococcus furiosus. Biochemistry 46:752-61

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