Assembly of phagocyte NADPH oxidase: A concerted binding process?

作者：Karimi Gilda; Levin Chantal Houee; Dagher Marie Claire; Baciou Laura; Bizouarn Tania^*

来源：Biochimica et Biophysica Acta-General Subjects, 2014, 1840(11): 3277-3283.

DOI：10.1016/j.bbagen.2014.07.022

• NADPH oxidase (Nox)
• Protein translocation
• Cell free system
• Arachidonic acid activation
• Neutrophil

Background: The phagocyte NADPH-oxidase is a multicomponent enzyme that generates superoxide anions. It comprises a membrane redox component flavocytochrome b(558) 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 spatiotemporal 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 b(558). 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.
Conclusion and general significance: 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.

• 出版日期2014-11