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Cryptic Rac-binding and p21(Cdc42Hs/Rac)-activated kinase phosphorylation sites of NADPH oxidase component p67(phox)

S Ahmed1, E Prigmore, S Govind

  • 1Department of Neurochemistry, Institute of Neurology, 1 Wakefield Street, London WC1N 1PJ, Great Britain.

The Journal of Biological Chemistry
|June 23, 1998
PubMed

Insights

Rac1 and Rac2 GTPases bind to a specific region of p67(phox), influencing NADPH oxidase activity. Deletions in p67(phox) enhance this binding and PAK phosphorylation, suggesting cryptic regulatory sites.

Area of Science:

  • Molecular Biology
  • Cell Signaling
  • Biochemistry

Background:

  • Rac1 and Rac2 GTPases are crucial for cell morphology, growth, and differentiation.
  • These GTPases are essential for superoxide production by the NADPH oxidase complex.
  • Previous studies showed Rac1 directly binds p67(phox).

Purpose of the Study:

  • To further characterize the interaction between Rac1/2 and p67(phox).
  • To identify specific binding sites and regulatory mechanisms controlling this interaction.
  • To investigate the role of p67(phox) phosphorylation by PAK in Rac signaling.

Main Methods:

  • Binding assays to map Rac1/2 interaction sites on p67(phox).
  • Site-directed mutagenesis and deletion analysis of p67(phox).
  • Investigating the effect of p67(phox) modifications on Rac1 binding and PAK phosphorylation.

Main Results:

  • Rac1 and Rac2 bind to amino acid residues 170-199 of p67(phox).
  • The N-terminus of p67(phox) (1-192) acts as a Rac signaling inhibitor.
  • Deletion of C-terminal domains (193-526), SH3 domain (470-526), or polyproline motif (226-236) increased Rac1 binding ~8-fold.
  • PAK phosphorylation of p67(phox) near the Rac-binding site is enhanced by C-terminal deletions.

Conclusions:

  • Specific regions of p67(phox) modulate Rac1/2 binding affinity.
  • Cryptic Rac-binding sites and PAK phosphorylation sites in p67(phox) likely regulate NADPH oxidase activity.
  • These findings provide insights into the intricate control of the NADPH oxidase complex.

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