Small-Angle X-ray Scattering (SAXS) Measurements of APOBEC3G Provide Structural Basis for Binding of Single-Stranded DNA and Processivity

Barzak F, Ryan T, Mohammadzadeh N, Harjes S, Kvach M, Kurup H, Krause K, Chelico L, Filichev V, Harjes E, Jameson G, Viruses 14(9):1974 (2022) DOI

SASDMY6 – single-stranded DNA dC->dU-editing enzyme APOBEC3G full length

DNA dC->dU-editing enzyme APOBEC-3G
MWexperimental 203 kDa
MWexpected 186 kDa
VPorod 350 nm3
log I(s) 4.41×10-3 4.41×10-4 4.41×10-5 4.41×10-6
DNA dC->dU-editing enzyme APOBEC-3G small angle scattering data  s, nm-1
ln I(s)
DNA dC->dU-editing enzyme APOBEC-3G Guinier plot ln 4.42×10-3 Rg: 4.2 nm 0 (4.2 nm)-2 s2
DNA dC->dU-editing enzyme APOBEC-3G Kratky plot 1.104 0 3 sRg
DNA dC->dU-editing enzyme APOBEC-3G pair distance distribution function Rg: 4.2 nm 0 Dmax: 13.3 nm

Data validation

Fits and models

log I(s)
 s, nm-1
DNA dC->dU-editing enzyme APOBEC-3G DAMFILT model

Synchrotron SAXS data from solutions of single-stranded DNA dC->dU-editing enzyme APOBEC3G in 50 mM phosphate pH 6.0, 200 mM NaCl, 2 mM β-mercaptoethanol (β-ME), 5% glycerol, 200 µM Na2-EDTA 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 and at a wavelength of λ = 0.10332 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). In-line size-exclusion chromatography (SEC) SAS was employed using a sheath-flow cell. The SEC parameters were as follows: A 100.00 μl sample at 1.3 mg/ml was injected at a 0.20 ml/min flow rate onto a GE Superdex 75 Increase 10/300 column at 25°C. 1000 successive 1 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 displayed bead model (damfilt) shows the bead-occupancy and volume corrected spatial representation of the protein calculated from the alignment of several individual models. An individual model-fit is displayed to the left.

DNA dC->dU-editing enzyme APOBEC-3G (A3G)
Mol. type   Protein
Organism   Homo sapiens
Olig. state   Tetramer
Mon. MW   46.4 kDa
UniProt   Q9HC16 (1-384)
Sequence   FASTA