On the structure and function of Escherichia coli YjhC: an oxidoreductase involved in bacterial sialic acid metabolism.

Horne CR, Kind L, Davies JS, Dobson RCJ, Proteins (2019) Europe PMC

SASDFZ3 – Escherichia coli YjhC

Escherichia coli YjhC
MWexperimental 85 kDa
MWexpected 86 kDa
VPorod 130 nm3
log I(s) 4.90×10-2 4.90×10-3 4.90×10-4 4.90×10-5
Escherichia coli YjhC small angle scattering data  s, nm-1
ln I(s)
Escherichia coli YjhC Guinier plot ln 4.90×10-2 Rg: 3.1 nm 0 (3.1 nm)-2 s2
(sRg)2I(s)/I(0)
Escherichia coli YjhC Kratky plot 1.104 0 3 sRg
p(r)
Escherichia coli YjhC pair distance distribution function Rg: 3.2 nm 0 Dmax: 10.7 nm

Data validation


Fits and models


log I(s)
 s, nm-1
Escherichia coli YjhC PDB model

Synchrotron SAXS data of the oxidoreductase YjhC from Escherichia coli in 20 mM Tris, 150 mM NaCl, 0.1 % (w/v) sodium azide, 5 % (v/v) glycerol, pH 8 were collected on the SAXS/WAXS beam line at the Australian Synchrotron (Melbourne, Australia) using a PILATUS 1M detector at a sample-detector distance of 1.6 m, providing a q range of 0.06-5 nm-1 and at a wavelength of λ = 0.10332 nm (l(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). Coflow size-exclusion chromatography-SAXS (SEC-SAXS) was used to minimize sample dilution and maximize signal to noise (Kirby et al., 2016). 80 µl of purified YjhC protein at 8 mg/ml was injected onto an Superdex S200 Increase 5/150 GL (GE Healthcare) at a flow rate of 0.45 ml/min. Scattering data was collected in 1 second exposures over a total of 400 frames, using a 1.5 mm glass capillary, at 12 °C. 2D intensity plots were radially averaged, normalised to sample transmission, and background subtracted using the Scatterbrain software package (Australian Synchrotron). Analysis of the scattering data was performed using the ATSAS software package (version 2.8.4). PrimusQT was used to perform the Guinier analysis, Kratky analysis, and to generate the pairwise distribution function P(r), calculated using an indirect Fourier transform using GNOM. The molecular mass of the samples was estimated using the SAXS-MoW2 package (Fischer et al., 2010), while CRYSOL was used to evaluate the experimental solution scattering with the calculated scattering from the atomic coordinates of the high-resolution structure of YjhC.

Escherichia coli YjhC (YjhC)
Mol. type   Protein
Organism   Escherichia coli (strain K12)
Olig. state   Dimer
Mon. MW   43.0 kDa
Sequence   FASTA
 
PDB code   6O15