SAXS data were collected from a solution of human deglycosylated alpha-2-macroglobulin treated with bovine pancreatic trypsin in a 20 mM HEPES, 150 mM NaCl, pH 7.4, buffer (I(s) vs s, where s = 4π sin θ/λ; 2θ is the scattering angle; λ = 0.134 nm). Each alpha-2-macroglobulin tetramer contains two bovine trypsin molecules, which results in a total molecular mass of 689 kDa for the alpha-2-macroglobulin tetramer of 161 kDa and two trypsins of 23 kDa. The data were collected at the in-house Bruker AXS NanoSTAR instrument with an Excillum Ga Metal Jet X-ray source at Aarhus University. The instrument has a homebuilt scatterless pinhole in front of the sample and is equipped with a Bruker AXS Vantec2000 detector. Data were collected for 1800 s for the sample and buffer, respectively, and the data were normalized to absolute scale using the scattering from pure water. The structure was modelled using home-written software, which performs rigid body refinement on the structure with P222 symmetry for alpha-2-macroglobulin and no symmetry for the two trypsins. The program takes into account a hydration layer.
|
|