A round-robin approach provides a detailed assessment of biomolecular small-angle scattering data reproducibility and yields consensus curves for benchmarking

Trewhella J, Vachette P, Bierma J, Blanchet C, Brookes E, Chakravarthy S, Chatzimagas L, Cleveland T, Cowieson N, Crossett B, Duff A, Franke D, Gabel F, Gillilan R, Graewert M, Grishaev A, Guss J, Hammel M, Hopkins J, Huang Q, Hub J, Hura G, Irving T, Jeffries C, Jeong C, Kirby N, Krueger S, Martel A, Matsui T, Li N, Pérez J, Porcar L, Prangé T, Rajkovic I, Rocco M, Rosenberg D, Ryan T, Seifert S, Sekiguchi H, Svergun D, Teixeira S, Thureau A, Weiss T, Whitten A, Wood K, Zuo X, Acta Crystallographica Section D Structural Biology 78(11) (2022) DOI

SASDPR4 – Consensus SAXS Profile - Xylose Isomerase

Xylose isomerase
MWexperimental 173 kDa
MWexpected 173 kDa
VPorod 243 nm3
log I(s) 1.05×100 1.05×10-1 1.05×10-2 1.05×10-3
Xylose isomerase small angle scattering data  s, nm-1
ln I(s)
Xylose isomerase Guinier plot ln 1.06×100 Rg: 3.3 nm 0 (3.3 nm)-2 s2
(sRg)2I(s)/I(0)
Xylose isomerase Kratky plot 1.104 0 √3 sRg
p(r)
Xylose isomerase pair distance distribution function Rg: 3.3 nm 0 Dmax: 10.1 nm

Data validation

Fits and models

log I(s)
 s, nm-1
Xylose isomerase DAMMIN model
log I(s)
 s, nm-1
Xylose isomerase PDB (PROTEIN DATA BANK) model
log I(s)
 s, nm-1
Xylose isomerase PDB (PROTEIN DATA BANK) model
log I(s)
 s, nm-1
Xylose isomerase PDB (PROTEIN DATA BANK) model
log I(s)
 s, nm-1
Xylose isomerase PDB (PROTEIN DATA BANK) model
The consensus SAXS profile for xylose isomerase was generated using the datcombine tool (ATSAS 3.1.0) with outlier-filter applied. The data input to datcombine were fourteen scattering profiles made up of pure SEC-SAXS (5), pure batch SAXS (6) 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 50 mM Tris, pH 7.5, 100 mM NaCl, 1 mM MgCl2. Protein concentrations for batch measurements ranged from 0.5 - 21.5 mg/mL, and all batch data > 1 mg/mL were merged with either SEC-SAXS or lower concentration data demonstrated to be free of interparticle interference, which is known to be significant for xylose isomerase concentrations > 1.0 mg/mL. The xylose isomerase atomistic model for CRYSOL, Pepsi-SAXS and FoXS calculations was derived from PDB ID 1MNZ tetramer with an N-terminal Met added using PyMol, and small-molecule crystallisation agents removed. Custom WAXSiS calculations (with Gromacs software) used the same coordinates and added explicit waters and appropriate number of ions for the MD calculation.

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
Xylose isomerase
Mol. type   Protein
Organism   Streptomyces rubiginosus
Olig. state   Tetramer
Mon. MW   43.2 kDa
 
UniProt   P24300 (1-388)
Sequence   FASTA
 
PDB ID   1MNZ
 
PDB ID   1MNZ