Synchrotron SAXS data from solutions of the ToxRp/ToxSp complex bound to bile salt in 50 mM Na2HPO4, 300 mM NaCl, 3% glycerol, pH 8 were collected on the EMBL P12 beam line at PETRA III (DESY, Hamburg, Germany) using a Pilatus 6M detector at a sample-detector distance of 3 m and at a wavelength of λ = 0.124 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). In-line size-exclusion chromatography (SEC) SAS was employed. The SEC parameters were as follows: A 100.00 μl sample at 10 mg/ml was injected at a 0.30 ml/min flow rate onto a GE Superdex 75 Increase 5/150 column at 20°C. 1800 successive 0.500 second frames were collected. The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted.
There is more than one bile molecule bound to the complex. Despite the use of SEC, it seems like the sample is slightly polydisperse with bile molecules weakly attaching to the surface of the ToxRSp complex (confirmed with a number of other techniques). Bile salt: https://pubchem.ncbi.nlm.nih.gov/compound/Sodium-cholate-hydrate
|
|