Tetratricopeptide repeat (TPR) motifs of p67(phox) participate in interaction with the small GTPase Rac and activation of the phagocyte NADPH oxidase.
The small GTPase Rac functions as a molecular switch in several important cellular events including cytoskeletal reorganization and activation of the phagocyte NADPH oxidase, the latter of which leads to production of superoxide, a precursor of microbicidal oxidants. During formation of the active oxidase complex at the membrane, the GTP-bound Rac appears to interact with the N-terminal region of p67(phox), another indispensable activator that translocates from the cytosol upon phagocyte stimulation. Here we show that the p67(phox) N terminus lacks the CRIB motif, a well known Rac target, but contains four tetratricopeptide repeat (TPR) motifs with highly alpha-helical structure. Disruption of any of the N-terminal three TPRs, but the last one, results in defective interaction with Rac, while all the four are required for the NADPH oxidase activation. We also find that Arg-102 in the third repeat is likely involved in binding to Rac via an ionic interaction, and that replacement of this residue with Glu completely abrogates the capability of activating the oxidase both in vivo and in vitro. Thus the TPR motifs of p67(phox) are packed to function as a Rac target, thereby playing a crucial role in the active oxidase complex formation.
Koga
H
H
Department of Molecular and Structural Biology, Kyushu University Graduate School of Medical Science, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
Terasawa
H
H
Nunoi
H
H
Takeshige
K
K
Inagaki
F
F
Sumimoto
H
H
D002942:Circular Dichroism; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D020014:K562 Cells; D009682:Magnetic Resonance Spectroscopy; D016296:Mutagenesis; D019255:NADPH Oxidases / Q000378:metabolism*; D009504:Neutrophils / Q000201:enzymology*; D010586:Phagocytes / Q000201:enzymology; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000235:genetics; D012091:Repetitive Sequences, Nucleic Acid; D016415:Sequence Alignment; D014162:Transfection; D015003:Yeasts / Q000235:genetics; D020744:rac GTP-Binding Proteins
D010750:Phosphoproteins; D011994:Recombinant Proteins; C079789:neutrophil cytosol factor 67K; D019255:NADPH Oxidases; D019204:GTP-Binding Proteins; D020744:rac GTP-Binding Proteins