The aim of this research proposal is to gain a clearer understanding of the mechanisms for the induction, synthesis and turnover of the fumarate reductase complex in Escherichia coli. This oxido-reductase enzyme complex is membrane bound and contains both flavoprotein and non-heme iron centers. Synthesis of the four individual polypeptides of the fumarate reductase complex occurs only under anaerobic growth conditions when fumarate is available for use as a terminal electron acceptor. Synthesis of the complex is repressed when oxygen or nitrate is present. Little is known about this induction and/or repression phenomenon at either the genetic or the molecular level. We plan to examine the mechanisms for the transcriptional regulation of the four genes encoding the individual polypeptides of the membrane bound fumarate reductase complex using recombinant DNA techniques. Mutations resulting in altered fumarate reductase expression as well as mutations in the structural genes for fumarate reductase will be generated to increase our understanding of both the regulatory phenomenon and biochemistry of the complex. The in vivo expression and turnover of the fumarate reductase complex under inducing and non inducing conditions will also be examined using gene fusion and immunological techniques. The fumarate, nitrate and FNR regulator proteins affecting frd expression will be studied using genetic and molecular approaches. Experiments directed towards understanding the activation of fumarate reductase expression by the positive regulatory protein, FNR, will be undertaken. The transcriptional regulation of the fnr gene encoding FNR will be examined to locate the promoter and associated regulatory control regions. We believe that fumarate reductase is an excellent model system to examine the induction and regulation of genes whose products are required under anaerobic growth conditions. Little is known about how these genes are induced or repressed in response to fumarate, nitrate and oxygen. Although the study of fumarate reductase in E. coli is interesting in its own right, this system may also serve as a useful model for understanding facultative metabolism in other enteric bacteria.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI021678-02
Application #
3131918
Study Section
Microbial Physiology and Genetics Subcommittee 2 (MBC)
Project Start
1986-09-01
Project End
1989-08-31
Budget Start
1987-09-01
Budget End
1988-08-31
Support Year
2
Fiscal Year
1987
Total Cost
Indirect Cost
Name
University of California Los Angeles
Department
Type
Schools of Arts and Sciences
DUNS #
119132785
City
Los Angeles
State
CA
Country
United States
Zip Code
90095
Katsir, Galit; Jarvis, Michael; Phillips, Martin et al. (2015) The Escherichia coli NarL receiver domain regulates transcription through promoter specific functions. BMC Microbiol 15:174
McInerney, Michael J; Sieber, Jessica R; Gunsalus, Robert P (2009) Syntrophy in anaerobic global carbon cycles. Curr Opin Biotechnol 20:623-32
Bose, Jeffrey L; Kim, Unmi; Bartkowski, Wojciech et al. (2007) Bioluminescence in Vibrio fischeri is controlled by the redox-responsive regulator ArcA. Mol Microbiol 65:538-53
Maris, Ann E; Kaczor-Grzeskowiak, Maria; Ma, Zhongcai et al. (2005) Primary and secondary modes of DNA recognition by the NarL two-component response regulator. Biochemistry 44:14538-52
Salmon, Kirsty A; Hung, She-pin; Steffen, Nicholas R et al. (2005) Global gene expression profiling in Escherichia coli K12: effects of oxygen availability and ArcA. J Biol Chem 280:15084-96
Rothery, Richard A; Seime, Andrea M; Spiers, A-M Caroline et al. (2005) Defining the Q-site of Escherichia coli fumarate reductase by site-directed mutagenesis, fluorescence quench titrations and EPR spectroscopy. FEBS J 272:313-26
Wang, Henian; Gunsalus, Robert P (2003) Coordinate regulation of the Escherichia coli formate dehydrogenase fdnGHI and fdhF genes in response to nitrate, nitrite, and formate: roles for NarL and NarP. J Bacteriol 185:5076-85
Salmon, Kirsty; Hung, She-pin; Mekjian, Kathy et al. (2003) Global gene expression profiling in Escherichia coli K12. The effects of oxygen availability and FNR. J Biol Chem 278:29837-55
Zhang, Jeffrey H; Xiao, Gaoping; Gunsalus, Robert P et al. (2003) Phosphorylation triggers domain separation in the DNA binding response regulator NarL. Biochemistry 42:2552-9
McNicholas, Paul M; Gunsalus, Robert P (2002) The molybdate-responsive Escherichia coli ModE transcriptional regulator coordinates periplasmic nitrate reductase (napFDAGHBC) operon expression with nitrate and molybdate availability. J Bacteriol 184:3253-9

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