Abstract

The leukocyte NADPH oxidase is the enzyme responsible for superoxide production by phagocytes and B lymphocytes. It is dormant in resting cells but comes to life when the cells are activated. Studies of oxidase activation in cell-free systems have used anionic detergents as activating agents, but oxidase activation in whole cells is accomplished by phosphorylation. As an extension of a study from another group, a kinase dependent cell-free activating system has been developed that will be used to study oxidase activation by what is probably the physiological route. Therefore, one aim of the present work is to characterize the kinase-dependent cell-free activating system, particularly with respect to effects on the membrane. The other aim concerns the further characterization of p67phox, an NADPH-binding oxidase component. The inactivation of p67phox by NADPH dialdehyde, an affinity label, will be studied in detail. In addition, p67phox will be purified in quantity, crystallized in the absence and presence of NADPH and analyzed by X-ray crystallography.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
2R01AI024227-14
Application #
2503938
Study Section
Hematology Subcommittee 2 (HEM)
Project Start
1986-03-01
Project End
2003-08-31
Budget Start
1998-09-01
Budget End
1999-08-31
Support Year
14
Fiscal Year
1998
Total Cost
Indirect Cost

  Institution

Name
Scripps Research Institute
Department
Type
DUNS #
City
La Jolla
State
CA
Country
United States
Zip Code
92037

  Publications

Johnson, Jennifer L; Brzezinska, Agnieszka A; Tolmachova, Tanya et al. (2010) Rab27a and Rab27b regulate neutrophil azurophilic granule exocytosis and NADPH oxidase activity by independent mechanisms. Traffic 11:533-47
Brzezinska, Agnieszka A; Johnson, Jennifer L; Munafo, Daniela B et al. (2009) Signalling mechanisms for Toll-like receptor-activated neutrophil exocytosis: key roles for interleukin-1-receptor-associated kinase-4 and phosphatidylinositol 3-kinase but not Toll/IL-1 receptor (TIR) domain-containing adaptor inducing IFN-beta (TRIF). Immunology 127:386-97
Munafo, Daniela B; Johnson, Jennifer L; Brzezinska, Agnieszka A et al. (2009) DNase I inhibits a late phase of reactive oxygen species production in neutrophils. J Innate Immun 1:527-42
Brzezinska, Agnieszka A; Johnson, Jennifer L; Munafo, Daniela B et al. (2008) The Rab27a effectors JFC1/Slp1 and Munc13-4 regulate exocytosis of neutrophil granules. Traffic 9:2151-64
Catz, Sergio D (2008) Characterization of Rab27a and JFC1 as constituents of the secretory machinery of prostate-specific antigen in prostate carcinoma cells. Methods Enzymol 438:25-40
Munafo, Daniela B; Johnson, Jennifer L; Ellis, Beverly A et al. (2007) Rab27a is a key component of the secretory machinery of azurophilic granules in granulocytes. Biochem J 402:229-39
Pacquelet, Sandrine; Johnson, Jennifer L; Ellis, Beverly A et al. (2007) Cross-talk between IRAK-4 and the NADPH oxidase. Biochem J 403:451-61
DeLano, Frank A; Parks, Dale A; Ruedi, Julie M et al. (2006) Microvascular display of xanthine oxidase and NADPH oxidase in the spontaneously hypertensive rat. Microcirculation 13:551-66
Johnson, Jennifer L; Ellis, Beverly A; Munafo, Daniela B et al. (2006) Gene transfer and expression in human neutrophils. The phox homology domain of p47phox translocates to the plasma membrane but not to the membrane of mature phagosomes. BMC Immunol 7:28
Johnson, Jennifer L; Ellis, Beverly A; Noack, Deborah et al. (2005) The Rab27a-binding protein, JFC1, regulates androgen-dependent secretion of prostate-specific antigen and prostatic-specific acid phosphatase. Biochem J 391:699-710

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