Quantitative live-cell imaging and 3D modeling reveal critical functional features in the cytosolic complex of phagocyte NADPH oxidase.

Ziegler CS, Bouchab L, Tramier M, Durand D, Fieschi F, Dupré-Crochet S, Mérola F, Nüße O, Erard M, J Biol Chem (2019) Europe PMC

SASDEK3 – The neutrophil cytosol factor 1 (p47phox) subunit of phagocyte NADPH oxidase

Neutrophil cytosol factor 1
MWI(0) 44 kDa
MWexpected 46 kDa
VPorod 77 nm3
log I(s) 3.27×10-2 3.27×10-3 3.27×10-4 3.27×10-5
Neutrophil cytosol factor 1 small angle scattering data  s, nm-1
ln I(s)
Neutrophil cytosol factor 1 Guinier plot ln 3.27×10-2 Rg: 3.2 nm 0 (3.2 nm)-2 s2
Neutrophil cytosol factor 1 Kratky plot 1.104 0 3 sRg
Neutrophil cytosol factor 1 pair distance distribution function Rg: 3.3 nm 0 Dmax: 12.5 nm

Data validation

Fits and models

log I(s)
 s, nm-1
Neutrophil cytosol factor 1 CUSTOM IN-HOUSE model

Synchrotron SAXS data from solutions of the p47phox subunit of phagocyte NADPH oxidase in 50 mM HEPES 50, 100 mM NaCl, 1 mM EDTA, 2 mM DTT, 5% glycerol, pH 7.5 were collected on the SWING beam line SOLEIL (Saint-Aubin, France) using a CCD AVIEX PCCD170170 detector at a sample-detector distance of 1.8 m and at a wavelength of λ = 0.1 nm (l(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). Samples were studied at different concentrations ranging from 1.0 to 4.0 mg/ml measured at 10°C. 20 successive 2 second frames were collected. SAXS data were normalized to the intensity of the incident beam, averaged and background subtracted using the program package PRIMUS. Intensities were put on an absolute scale (cm-1) using water scattering. The final pattern used for fitting was obtained by extrapolating to infinite dilution of the set of curves recorded at the different concentrations. Molecular mass was obtained using the bayesian inference approach proposed in PRIMUS/qt, which combines four concentration independent MM estimators: M=47.7 kDa within the credibility interval [45.2-50.3].

Neutrophil cytosol factor 1 (p47phox)
Mol. type   Protein
Organism   Homo sapiens
Olig. state   Monomer
Mon. MW   46.4 kDa
UniProt   P14598 (1-390)
Sequence   FASTA