Biomolecular Small-Angle Scattering Data Reproducibility and Consensus Curves for Benchmarking

Jill Trewhella.

SASDPQ4 – Consensus SAXS Profile - Urate Oxidase

Uricase
MWexperimental 137 kDa
MWexpected 136 kDa
VPorod 220 nm3
log I(s) 1.12×100 1.12×10-1 1.12×10-2 1.12×10-3
Uricase small angle scattering data  s, nm-1
ln I(s)
Uricase Guinier plot ln 1.13×100 Rg: 3.2 nm 0 (3.2 nm)-2 s2
(sRg)2I(s)/I(0)
Uricase Kratky plot 1.104 0 3 sRg
p(r)
Uricase pair distance distribution function Rg: 3.2 nm 0 Dmax: 9.2 nm

Data validation


Fits and models


log I(s)
 s, nm-1
Uricase DAMMIN model

log I(s)
 s, nm-1
Uricase PDB (PROTEIN DATA BANK) model

log I(s)
 s, nm-1
Uricase PDB (PROTEIN DATA BANK) model

log I(s)
 s, nm-1
Uricase PDB (PROTEIN DATA BANK) model

log I(s)
 s, nm-1
Uricase PDB (PROTEIN DATA BANK) model

The consensus SAXS profile for urate oxidase was generated by the datcombine tool (ATSAS 3.1.0) with both outlier- and error-filters applied. The data input to datcombine were eleven scattering profiles made up of pure SEC-SAXS (6), pure batch SAXS (2) and merged SEC-SAXS-batch SAXS (3) data. All contributing data were independent measurements, and no individual measurement was represented more than once in the contributing scattering profile set. The buffer for substantial majority of the contributing data was 100 mM Tris, pH 8.0, 150 mM NaCl. Protein concentrations for batch measurements ranged from 1 - 7 mg/mL, and all batch data that showed evidence of aggregation were merged with SEC-SAXS or lower concentration data to remove any influence from aggregate. The urate oxidase atomistic model for CRYSOL, Pepsi-SAXS, and FoXS calculations was PDB ID 3L8W tetramer with small-molecule crystallisation agents removed, and a Modloop generated C-terminal sequence SLKSKL added to reflect the sequence for the protein as measured using SAXS. Custom WAXSiS calculations (with Gromacs software) used the same coordinates and added explicit waters and appropriate number of ions for the MD calculations.

The data input to datcombine are made available for download in the associated zip file. Model fits are shown in order (top to bottom): DAMMIN, CRYSOL, Pepsi-SAXS, FoXS, and custom WAXSiS. The unusually good statistics for the consensus SAXS data generally give rise to large χ-square values for the model fits.

Tags: benchmark
Uricase
Mol. type   Protein
Organism   Aspergillus flavus
Olig. state   Tetramer
Mon. MW   34.1 kDa
 
UniProt   Q00511 (2-302)
Sequence   FASTA
 
PDB ID   3L8W
 
PDB ID   3L8W