Modeling the ComD/ComE/comcde interaction network using small angle X-ray scattering.

Sanchez D, Boudes M, van Tilbeurgh H, Durand D, Quevillon-Cheruel S, FEBS J 282(8):1538-53 (2015) Europe PMC

SASDAB7 – Complex ComE-comcde

comcde
Response regulator
MWexperimental 83 kDa
MWexpected 85 kDa
VPorod 122 nm3
log I(s) 5.52×10-2 5.52×10-3 5.52×10-4 5.52×10-5
comcde Response regulator small angle scattering data  s, nm-1
ln I(s)
comcde Response regulator Guinier plot ln 5.53×10-2 Rg: 3.4 nm 0 (3.4 nm)-2 s2
(sRg)2I(s)/I(0)
comcde Response regulator Kratky plot 1.104 0 √3 sRg
p(r)
comcde Response regulator pair distance distribution function Rg: 3.4 nm 0 Dmax: 10.7 nm

Data validation

Fits and models

log I(s)
 s, nm-1
comcde Response regulator OTHER model
X-ray synchrotron radiation scattering data from solutions of complex ComE-comcde in 50 mM MES 500 mM NaCl, 5% (vol/vol) glycerol, 5 mM β-mercaptoethanol were collected on the SWING beamline on the storage ring SOLEIL (Saint-Aubin, France) using a CCD AVIEX detector (s = 4π sin θ/λ, where 2θ is the scattering angle). The sample was loaded on a SE-HPLC column on line with the SAXS measuring cell. The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted. The scattered intensities were displayed on an absolute scale (cm-1) using the scattering of water. Frames were examined individually and 6 identical frames were averaged and further processed. The corresponding concentration was 0.8 g/L.

Use of SAXS to describe the interaction between the phosphorylated mimetic mutant ComED58E of the response regulator ComE from S. pneumonia and its promoter region comcde. The complex is constituted of one dimer of ComE and one monomer of comcde. The SAXS data were collected directly after elution through an on-line size-exclusion high-performance liquid chromatography (SEC-HPLC) column. The molecular weight is not estimated from I(0) because the concentration of the complex cannot be accurately determined. It is obtained from the macromolecule volume using the method developed by Craievich’s team (the SAXS Mow program).

comcde
Mol. type   DNA
Organism   Streptococcus pneumoniae
Olig. state   Other
Mon. MW   23.5 kDa
Sequence   FASTA
 
Response regulator (ComE)
Mol. type   Protein
Organism   Streptococcus pneumoniae
Olig. state   Dimer
Mon. MW   30.7 kDa
 
UniProt   Q8DMW5 (1-250)
Sequence   FASTA