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Synchrotron SAXS data from solutions of gelsolin in 20 mM HEPES, 100 mM NaCl, 1 mM EDTA, pH 7.4 were collected on the B21 beam line at the Diamond Light Source (Didcot, UK) using a Eiger 4M detector at a sample-detector distance of 3.5 m and at a wavelength of λ = 0.1 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). One solute concentration of 2.01 mg/ml was measured at 25°C. 20 successive 1 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.
Human plasma isoform of GSN devoid of the signal peptide (mature form) was produced in E. coli SHuffle® cells (New England Biolabs) upon addition of 0.5 mM IPTG and incubation for 16 hours at 18 °C. Cells were lysed in a Basic Z Bench top (Constant Systems Limited, U.K.) at 25 kPSI, and the clarified extract passed through a HisTrap HP column (all chromatographic media from GE-Healthcare). Further polishing was obtained by anion-exchange (Resource Q) followed by size-exclusion chromatography (HiLoad 16/600 Superdex 200).
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s, nm-1
