The Active N-terminal Region of p67phox

Sylvestre Grizot, Franck Fieschi, Marie-Claire Dagher, Eva Pebay-Peyroula
2001 Journal of Biological Chemistry  
Upon activation, the NADPH oxidase from neutrophils produces superoxide anions in response to microbial infection. This enzymatic complex is activated by association of its cytosolic factors p67 phox , p47 phox , and the small G protein Rac with a membrane-associated flavocytochrome b 558 . Here we report the crystal structure of the active N-terminal fragment of p67 phox at 1.8 Å resolution, as well as functional studies of p67 phox mutants. This N-terminal region (residues 1-213) consists
more » ... ly of four TPR (tetratricopeptide repeat) motifs in which the C terminus folds back into a hydrophobic groove formed by the TPR domain. The structure is very similar to that of the inactive truncated form of p67 phox bound to the small G protein Rac previously reported, but differs by the presence of a short C-terminal helix (residues 187-193) that might be part of the activation domain. All p67 phox mutants responsible for Chronic Granulomatous Disease (CGD), a severe defect of NADPH oxidase function, are localized in the N-terminal region. We investigated two CGD mutations, G78E and A128V. Surprisingly, the A128V CGD mutant is able to fully activate the NADPH oxidase in vitro at 25°C. However, this point mutation represents a temperaturesensitive defect in p67 phox that explains its phenotype at physiological temperature. The NADPH oxidase of phagocytic cells is responsible for the production of microbicidal superoxide anions. This enzymatic complex is activated at the onset of phagocytosis by association of its cytosolic factors p67 phox , p47 phox , and the small G protein Rac with a membrane-associated flavocytochrome b 558 (1). Mutations in any of these components except Rac can lead to a severe immune defect known as Chronic Granulomatous Disease (CGD). 1 The cytosolic factors p47 phox , p40 phox , and p67 phox are modular proteins comprising a set of structural domains such as SH3 domains, proline-rich regions, and TPR motifs (2),
doi:10.1074/jbc.m100893200 pmid:11262407 fatcat:hfsrmq3ojbgidlzr62xzfvzyvy