title: Assembly of phagocyte NADPH oxidase: A concerted binding process?
abstract: BACKGROUND
  The phagocyte NADPH-oxidase is a multicomponent enzyme that generates superoxide anions. It comprises a membrane redox component flavocytochrome b558 and four cytosolic proteins (p67(phox), p47(phox), p40(phox) and Rac) that must assemble to produce an active system. In this work we focused on the spatio-temporal control of the activation process of phagocyte NADPH oxidase.
  
  
  METHODS
  A wide range of techniques including fast kinetics with a stopped-flow apparatus and various combinations of the activating factors was used to test the order of assembly and the role of the p47(phox)-p67(phox) complex.
  
  
  RESULTS
  The data presented here are consistent with the absence of a catalytic role of the p47(phox)-p67(phox) interacting state and support the idea of independent binding sites for the cytosolic proteins on the flavocytochrome b558 allowing random binding order. However, the formation of the active complex appears to involve a synergistic process of binding of the activated cytosolic subunits to cytochrome b558. All partners should be in the vicinity for optimal assembly, a delay or the absence of one of the partners in this process seems to lead to a decrease in the efficiency of the catalytic core.
  
  
  CONCLUSIONS
  The activation and assembly of the NADPH oxidase components have to be achieved simultaneously for the formation of an efficient and optimal enzyme complex. This mechanism appears to be incompatible with continuous fast exchanges of the cytosolic proteins during the production of superoxide ion in the phagosome.
authors.affiliation: Laboratoire de Chimie Physique, Université Paris Sud, UMR8000, CNRS, Orsay F-91405, France.
authors.forename: Gilda
authors.initials: G
authors.lastname: Karimi
authors.affiliation: Laboratoire de Chimie Physique, Université Paris Sud, UMR8000, CNRS, Orsay F-91405, France.
authors.forename: Chantal
authors.initials: C
authors.lastname: Houée Levin
authors.affiliation: Laboratoire TIMC-IMAG, Equipe TheREx, UMR5525, 38710 La Tronche, France.
authors.forename: Marie Claire
authors.initials: MC
authors.lastname: Dagher
authors.affiliation: Laboratoire de Chimie Physique, Université Paris Sud, UMR8000, CNRS, Orsay F-91405, France.
authors.forename: Laura
authors.initials: L
authors.lastname: Baciou
authors.affiliation: Laboratoire de Chimie Physique, Université Paris Sud, UMR8000, CNRS, Orsay F-91405, France; Laboratoire de Chimie Physique, bat 350, Université Paris Sud, 91405 Orsay cedex, France. Electronic address: tania.bizouarn@u-psud.fr.
authors.forename: Tania
authors.initials: T
authors.lastname: Bizouarn
keywords: Arachidonic acid activation; Cell free system; NADPH oxidase (Nox); Neutrophil; Protein translocation