Activity and structure of EcoKMcrA.

Czapinska H, Kowalska M, Zagorskaite E, Manakova E, Slyvka A, Xu SY, Siksnys V, Sasnauskas G, Bochtler M, Nucleic Acids Res 46(18):9829-9841 (2018) Europe PMC

SASDJW2 – Modification dependent EcoKMcrA restriction endonuclease

5-methylcytosine-specific restriction enzyme A
MWI(0) 61 kDa
MWexpected 65 kDa
VPorod 95 nm3
log I(s) 3.54×103 3.54×102 3.54×101 3.54×100
5-methylcytosine-specific restriction enzyme A small angle scattering data  s, nm-1
ln I(s)
5-methylcytosine-specific restriction enzyme A Guinier plot ln 3.55×103 Rg: 3.7 nm 0 (3.7 nm)-2 s2
(sRg)2I(s)/I(0)
5-methylcytosine-specific restriction enzyme A Kratky plot 1.104 0 √3 sRg
p(r)
5-methylcytosine-specific restriction enzyme A pair distance distribution function Rg: 3.8 nm 0 Dmax: 13 nm

Data validation

Fits and models

log I(s)
 s, nm-1
5-methylcytosine-specific restriction enzyme A PDB (PROTEIN DATA BANK) model
Synchrotron SAXS data from solutions of EcoKMcrA restriction endonuclease in 20 mM Tris–HCl pH 7.5, 200 mM KCl, 0.1 mM EDTA, 0.01% (w/v) sodium azide and 1 mM DTT, were collected on the EMBL P12 beam line at PETRA III (DESY, Hamburg, Germany) using a Pilatus 2M detector at a sample-detector distance of 3.1 m and at a wavelength of λ = 0.124 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). Solute concentrations ranging between 2 and 5.2 mg/ml were measured at 20°C. 20 successive 0.050 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 concentration were merged with the highest concentration high angle data to yield the final composite scattering curve.

Apo EcoKMcrA dimer

5-methylcytosine-specific restriction enzyme A (EcoKMcrA)
Mol. type   Protein
Organism   Escherichia coli
Olig. state   Dimer
Mon. MW   32.7 kDa
 
UniProt   P24200 (1-277)
Sequence   FASTA
 
PDB ID   6GHC