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

SASDEB5 – Bovine serum albumin from in-house SAXS

Bovine serum albumin
MWI(0) 64 kDa
MWexpected 66 kDa
VPorod 99 nm3
log I(s) 2.12×10-1 2.12×10-2 2.12×10-3 2.12×10-4
Bovine serum albumin small angle scattering data  s, nm-1
ln I(s)
Bovine serum albumin Guinier plot ln 2.12×10-1 Rg: 2.8 nm 0 (2.8 nm)-2 s2
(sRg)2I(s)/I(0)
Bovine serum albumin Kratky plot 1.104 0 √3 sRg
p(r)
Bovine serum albumin pair distance distribution function Rg: 2.8 nm 0 Dmax: 9.4 nm

Data validation

Fits and models

log I(s)
 s, nm-1
Bovine serum albumin PDB (PROTEIN DATA BANK) model
log I(s)
 s, nm-1
Bovine serum albumin DAMFILT model
SAXS data from solutions of Bovine serum albumin from in-house SAXS in 50 mM HEPES, pH 7.5 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 sample-detector distance of 0.7 m and at a wavelength of λ = 0.134 nm (l(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). One solute concentration of 4.60 mg/ml was measured at 25°C. One 60 second frame was collected. The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted.

The background corrected data was binned logarithmically, normalized by concentration and brought to absolute scale using water as a secondary standard.

Bovine serum albumin (BSA)
Mol. type   Protein
Organism   Bos taurus
Olig. state   Monomer
Mon. MW   66.4 kDa
 
UniProt   P02769 (25-607)
Sequence   FASTA
 
PDB ID   4F5S