The molecular basis of Rac-GTP action—promoting binding of p67phox to Nox2 by disengaging the β hairpin from downstream residues

Edna Bechor, Anat Zahavi, Yevgeny Berdichevsky, Edgar Pick*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

7 Scopus citations


p67phox fulfils a key role in the assembly/activation of the NADPH oxidase by direct interaction with Nox2. We proposed that Rac-GTP serves both as a carrier of p67phox to the membrane and an inducer of a conformational change enhancing its affinity for Nox2. This study provides evidence for the latter function: (i) oxidase activation was inhibited by p67phox peptides (106–120) and (181–195), corresponding to the β hairpin and to a downstream region engaged in intramolecular bonds with the β hairpin, respectively; (ii) deletion of residues 181–193 and point mutations Q115R or K181E resulted in selective binding of p67phox to Nox2 peptide (369–383); (iii) both deletion and point mutations led to a change in p67phox, expressed in increased apparent molecular weights; (iv) p67phox was bound to p67phox peptide (181–195) and to a cluster of peptides (residues 97–117), supporting the participation of selected residues within these sequences in intramolecular bonds; (v) p67phox failed to bind to Nox2 peptide (369–383), following interaction with Rac1-GTP, but a (p67phox-Rac1-GTP) chimera exhibited marked binding to the peptide, similar to that of p67phox deletion and point mutants; and (vi) size exclusion chromatography of the chimera revealed its partition in monomeric and polymeric forms, with binding to Nox2 peptide (369–383) restricted to polymers. The molecular basis of Rac-GTP action entails unmasking of a previously hidden Nox2-binding site in p67phox, following disengagement of the β hairpin from more C-terminal residues. The domain in Nox2 binding the “modified” p67phox comprises residues within the 369–383 sequence in the cytosolic dehydrogenase region.

Original languageEnglish
Pages (from-to)219-237
Number of pages19
JournalJournal of Leukocyte Biology
Issue number2
StatePublished - Aug 2021


FundersFunder number
Joseph and Shulamit Salomon Fund
Israel Science Foundation
Tel Aviv University


    • (p67-Rac) chimera
    • NADPH oxidase
    • conformational change
    • intramolecular bond
    • peptide-protein interaction
    • synthetic peptides


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