A conserved region between the TPR and activation domains of p67 phox participates in activation of the phagocyte NADPH oxidase

Yuichi Maehara, Kei Miyano, Satoru Yuzawa, Risa Akimoto, Ryu Takeya, Hideki Sumimoto

研究成果: Contribution to journalArticle査読

20 被引用数 (Scopus)

抄録

The phagocyte NADPH oxidase, dormant in resting cells, is activated during phagocytosis to produce superoxide, a precursor of microbicidal oxidants. The membrane-integrated protein gp91phox serves as the catalytic core, because it contains a complete electron-transporting apparatus from NADPH to molecular oxygen for superoxide production. Activation of gp91phox requires the cytosolic proteins p67phox, p47phox, and Rac (a small GTPase). p67phox, comprising 526 amino acids, moves upon cell stimulation to the membrane together with p47phox and there interacts with Rac; these processes are prerequisite for gp91phox activation. Here we show that a region of p67phox (amino acids 190-200) C-terminal to the Rac-binding domain is evolutionarily well conserved and participates in oxidase activation at a later stage in conjunction with an activation domain. Alanine substitution for Tyr-198, Leu-199, or Val-204 abrogates the ability of p67phox to support superoxide production by gp91phox-based oxidase as well as its related oxidases Nox1 and Nox3; the activation also involves other invariant residues such as Leu-193, Asp-197, and Gly-200. Intriguingly, replacement of Gln-192 by alanine or that of Tyr-198 by phenylalanine or tryptophan rather enhances superoxide production by gp91phox-based oxidase, suggesting a tuning role for these residues. Furthermore, the Y198A/V204A or L199A/V204A substitution leads to not only a complete loss of the activity of the reconstituted oxidase system but also a significant decrease in p67phox interaction with the gp91 phox NADPH-binding domain, although these mutations affect neither the protein integrity nor the Rac binding activity. Thus the extended activation domain of p67phox (amino acids 190-210) containing the D(Y/F)LGK motif plays an essential role in oxidase activation probably by interacting with gp91phox.

本文言語英語
ページ(範囲)31435-31445
ページ数11
ジャーナルJournal of Biological Chemistry
285
41
DOI
出版ステータス出版済み - 10 8 2010

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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