Structural and Mechanistic Insights into Caffeine Degradation by the Bacterial N-Demethylase Complex.

Kim JH, Kim BH, Brooks S, Kang SY, Summers RM, Song HK, J Mol Biol 431(19):3647-3661 (2019) Europe PMC

SASDFD7 – Pseudomonas putida CBB5 NdmAB complex - static

Methylxanthine N1-demethylase NdmA
Methylxanthine N3-demethylase NdmB
MWexperimental 271 kDa
MWexpected 256 kDa
log I(s) 4.02×101 4.02×100 4.02×10-1 4.02×10-2
Methylxanthine N1-demethylase NdmA Methylxanthine N3-demethylase NdmB small angle scattering data  s, nm-1
ln I(s)
Methylxanthine N1-demethylase NdmA Methylxanthine N3-demethylase NdmB Guinier plot ln 4.03×101 Rg: 4.5 nm 0 (4.5 nm)-2 s2
(sRg)2I(s)/I(0)
Methylxanthine N1-demethylase NdmA Methylxanthine N3-demethylase NdmB Kratky plot 1.104 0 3 sRg
p(r)
Methylxanthine N1-demethylase NdmA Methylxanthine N3-demethylase NdmB pair distance distribution function Rg: 4.3 nm 0 Dmax: 12.3 nm

Data validation


Fits and models


log I(s)
 s, nm-1
Methylxanthine N1-demethylase NdmA Methylxanthine N3-demethylase NdmB DAMMIF model

Synchrotron SAXS data from solutions of Pseudomonas putida CBB5 NdmAB complex - static in 20 mM HEPES 150 mM NaCl 2 mM TCEP 10% v/v glycerol, pH 7.5 were collected on the 4C beam line at the Pohang Accelerator Laboratory storage ring (Pohang, South Korea) using a Rayonix SX165 detector at a wavelength of λ = 0.0733 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). Solute concentrations ranging between 4.0 and 10.1 mg/ml were measured at 4°C. Six successive 5 second frames were collected. The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted. The low angle data collected at lower concentrations were extrapolated to infinite dilution and merged with the higher concentration data to yield the final composite scattering curve.

Methylxanthine N1-demethylase NdmA
Mol. type   Protein
Organism   Pseudomonas putida
Olig. state   Trimer
Mon. MW   42.3 kDa
 
UniProt   H9N289
Sequence   FASTA
 
Methylxanthine N3-demethylase NdmB
Mol. type   Protein
Organism   Pseudomonas putida
Olig. state   Trimer
Mon. MW   43.0 kDa
 
UniProt   H9N290
Sequence   FASTA