Synchrotron SAXS
data from solutions of
Fep1-rec-GATA
in
50 mM MOPS, 50 mM NaCl, pH 7
were collected
on the
BM29 beam line
at the ESRF storage ring
(Grenoble, France)
using a Pilatus 1M detector
at a sample-detector distance of 2.9 m and
at a wavelength of λ = 0.0999 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 40.00 μl sample
was injected at a 0.40 ml/min flow rate
onto a GE Superdex 75 Increase 5/150 column
at 20°C.
900 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.
The protein Fep1 was first reconstituted to saturate all the possible [2Fe2S] clusters, as described in the previous project.
Then it was purified and this purified Fep1-rec was incubated with a stoichiometric amount of double strand cognate GATA element.
The protein and the DNA were both 100 microM.
The complex was then loaded into a SEC column and the scattering of the elution solution was collected in continuous.
s, nm-1
