Abstract

The enzymology and regulation of neutrophil oxidative enzymes will be studied, with particular emphasis on several plasma membrane-associated components. In particular, we will study two enzyme systems: (1) the activatable NADPH-oxidase which is responsible for the superoxide generation of the oxidative burst, and which functions normally in bacterial killing by neutrophils, and (2) a newly discovered NADH-ferricyanide reductase of unknown function. The former is likely to consist minimally flavoprotein plus the recently discovered cytochrome b558. The prosthetic group of the latter is unknown. Defects in the components of the oxidase produce the inherited chronic granulomatous disease, while inappropriate activation of the system participates in the tissue damage seen in a variety of inflamatory diseases (e.g. shock lung, rheumatoid arthritis). Techniques for the study of these enzymes will include UV-visible absorbance spectrophotometry, liquid helium-temperature EPR (electron paramagnetic resonance), stopped flow spectrophotometry, and kinetic approaches.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI022809-03
Application #
3134351
Study Section
Physical Biochemistry Study Section (PB)
Project Start
1986-08-01
Project End
1989-09-29
Budget Start
1988-08-01
Budget End
1989-09-29
Support Year
3
Fiscal Year
1988
Total Cost
Indirect Cost

  Institution

Name
Emory University
Department
Type
Schools of Medicine
DUNS #
042250712
City
Atlanta
State
GA
Country
United States
Zip Code
30322

  Publications

Nisimoto, Y; Motalebi, S; Han, C H et al. (1999) The p67(phox) activation domain regulates electron flow from NADPH to flavin in flavocytochrome b(558). J Biol Chem 274:22999-3005
Han, C H; Freeman, J L; Lee, T et al. (1998) Regulation of the neutrophil respiratory burst oxidase. Identification of an activation domain in p67(phox). J Biol Chem 273:16663-8
Nisimoto, Y; Freeman, J L; Motalebi, S A et al. (1997) Rac binding to p67(phox). Structural basis for interactions of the Rac1 effector region and insert region with components of the respiratory burst oxidase. J Biol Chem 272:18834-41
Ogata, K; Nishimoto, N; Uhlinger, D J et al. (1996) Spermine suppresses the activation of human neutrophil NADPH oxidase in cell-free and semi-recombinant systems. Biochem J 313 ( Pt 2):549-54
Kreck, M L; Freeman, J L; Abo, A et al. (1996) Membrane association of Rac is required for high activity of the respiratory burst oxidase. Biochemistry 35:15683-92
Olson, S C; Lambeth, J D (1996) Biochemistry and cell biology of phospholipase D in human neutrophils. Chem Phys Lipids 80:3-19
Freeman, J L; Lambeth, J D (1996) NADPH oxidase activity is independent of p47phox in vitro. J Biol Chem 271:22578-82
Freeman, J L; Abo, A; Lambeth, J D (1996) Rac ""insert region"" is a novel effector region that is implicated in the activation of NADPH oxidase, but not PAK65. J Biol Chem 271:19794-801
Nisimoto, Y; Otsuka-Murakami, H; Lambeth, D J (1995) Reconstitution of flavin-depleted neutrophil flavocytochrome b558 with 8-mercapto-FAD and characterization of the flavin-reconstituted enzyme. J Biol Chem 270:16428-34
Uhlinger, D J; Tyagi, S R; Lambeth, J D (1995) On the mechanism of inhibition of the neutrophil respiratory burst oxidase by a peptide from the C-terminus of the large subunit of cytochrome b558. Biochemistry 34:524-7

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