Size-exclusion chromatography small-angle X-ray scattering of water soluble proteins on a laboratory instrument.

Bucciarelli S, Midtgaard SR, Nors Pedersen M, Skou S, Arleth L, Vestergaard B, J Appl Crystallogr 51(Pt 6):1623-1632 (2018) Europe PMC

SASDE55 – Apoferritin monomer from in-house SEC-SAXS

Ferritin light chain
MWexperimental 437 kDa
MWexpected 476 kDa
VPorod 685 nm3
log I(s) 2.24×100 2.24×10-1 2.24×10-2 2.24×10-3
Ferritin light chain small angle scattering data  s, nm-1
ln I(s)
Ferritin light chain Guinier plot ln 2.25×100 Rg: 5.1 nm 0 (5.1 nm)-2 s2
Ferritin light chain Kratky plot 1.104 0 3 sRg
Ferritin light chain pair distance distribution function Rg: 5.2 nm 0 Dmax: 12.3 nm

Data validation

Fits and models

log I(s)
 s, nm-1
Ferritin light chain PDB model

log I(s)
 s, nm-1
Ferritin light chain DAMFILT model

SAXS data from solutions of Apoferritin monomer from in-house SEC-SAXS in PBS, pH 7.4 were collected on a Xenocs BioXolver L instrument at the Copenhagen University, Department of Drug Design and Pharmacology (Copenhagen, Denmark) using a 20Hz Pilatus 300K detector at a wavelength of λ = 0.134 nm (l(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle).

The HAF sample (500 microlitres at 11.1 mg/ml) was injected onto a GE Healthcare S200 Increase 10/300 (24 ml) column at a flow rate of 0.1 ml/min 23°C. A total of 100 x 30 second SAXS data frames were recorded throughout the HAF elution at a sample-detector distance of 0.654 m and 110 x 30 second SAXS data frames at a sample-detector distance of 1.507 m. After background solvent corrections, SEC-SAXS data frames 65-75 and 66-76, respectively, corresponding to monomeric HAF, were scaled, averaged, binned logarithmically and merged to produce the data displayed in this entry. Additional files, including plots of the selected frames and of I(0), HPLC and exposure cell UV traces and Rg vs. time (determined by AUTORG) in both sample-detector settings are included in the additional files of the full entry zip archive.

Ferritin light chain (HAF)
Mol. type   Protein
Organism   Equus caballus
Olig. state   24-mer
Mon. MW   19.8 kDa
UniProt   P02791
Sequence   FASTA
PDB code   1IES