Mechanism of NanR gene repression and allosteric induction of bacterial sialic acid metabolism

Horne C, Venugopal H, Panjikar S, Henrickson A, Brookes E, North R, Murphy J, Friemann R, Griffin M, Ramm G, Demeler B, Dobson R, (2020) DOI

SASDHS9 – GntR-type transcriptional regulator NanR from Escherichia coli in the presence of N-acetylneuraminic acid

HTH-type transcriptional repressor NanR
MWexperimental 63 kDa
MWexpected 59 kDa
VPorod 105 nm3
log I(s) 3.23×10-2 3.23×10-3 3.23×10-4 3.23×10-5
HTH-type transcriptional repressor NanR small angle scattering data  s, nm-1
ln I(s)
HTH-type transcriptional repressor NanR Guinier plot ln 3.24×10-2 Rg: 3.1 nm 0 (3.1 nm)-2 s2
(sRg)2I(s)/I(0)
HTH-type transcriptional repressor NanR Kratky plot 1.104 0 √3 sRg
p(r)
HTH-type transcriptional repressor NanR pair distance distribution function Rg: 3.2 nm 0 Dmax: 10.1 nm

Data validation

Fits and models

log I(s)
 s, nm-1
HTH-type transcriptional repressor NanR CUSTOM IN-HOUSE model
Synchrotron SAXS data from solutions of NanR from Escherichia coli in 20 mM Tris-HCl (pH 8.0), 150 mM NaCl, 0.1 % (w/v) sodium azide, supplemented with 20 mM Neu5Ac were collected using co-flow size exclusion chromatography SAXS (SEC-SAXS) 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 and at a wavelength of λ = 0.10332 nm (l(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). Co-flow SEC-SAXS was performed at 12°C using the following parameters: Column: Superdex S200 Increase 5/150 GL (GE Healthcare); Flow rate: 0.45 mL/min; Sample injection concentration: 10 mg/mL; Injection volume: 70 μL. The data were collected through the SEC peak of the protein as a series of 1 second exposures. Each unsubtracted data frame was normalised to the intensity of the transmitted beam and radially averaged using the Scatterbrain software package (Australian Synchrotron). The scattering of an appropriate solvent-blank was subtracted. The resulting subtracted frames were scaled and averaged to generate the final SAXS profile displayed in this entry.

HTH-type transcriptional repressor NanR (NanR)
Mol. type   Protein
Organism   Escherichia coli
Olig. state   Dimer
Mon. MW   29.5 kDa
 
UniProt   P0A8W0 (1-263)
Sequence   FASTA