Prokaryotic Argonaute from Archaeoglobus fulgidus interacts with DNA as a homodimer.

Golovinas E, Rutkauskas D, Manakova E, Jankunec M, Silanskas A, Sasnauskas G, Zaremba M, Sci Rep 11(1):4518 (2021) Europe PMC

SASDH39 – Complex of Archaeoglobus fulgidus Argonaute with dsDNA (5'p-ATTGTACGTACAAT)

Piwi protein AF_1318
5'-phosphorilated 14-mer DNA oligoduplex
MWI(0) 100 kDa
MWexpected 110 kDa
VPorod 158 nm3
log I(s) 7.25×10-2 7.25×10-3 7.25×10-4 7.25×10-5
Piwi protein AF_1318 5'-phosphorilated 14-mer DNA oligoduplex small angle scattering data  s, nm-1
ln I(s)
Piwi protein AF_1318 5'-phosphorilated 14-mer DNA oligoduplex Guinier plot ln 7.25×10-2 Rg: 3.2 nm 0 (3.2 nm)-2 s2
(sRg)2I(s)/I(0)
Piwi protein AF_1318 5'-phosphorilated 14-mer DNA oligoduplex Kratky plot 1.104 0 √3 sRg
p(r)
Piwi protein AF_1318 5'-phosphorilated 14-mer DNA oligoduplex pair distance distribution function Rg: 3.2 nm 0 Dmax: 10.1 nm

Data validation

Fits and models

log I(s)
 s, nm-1
Piwi protein AF_1318 5'-phosphorilated 14-mer DNA oligoduplex PDB (PROTEIN DATA BANK) model
log I(s)
 s, nm-1
Piwi protein AF_1318 5'-phosphorilated 14-mer DNA oligoduplex PDB (PROTEIN DATA BANK) model
log I(s)
 s, nm-1
Piwi protein AF_1318 5'-phosphorilated 14-mer DNA oligoduplex DAMMIN model
Synchrotron SAXS data from solutions of Argonaute bound to dsDNA (5'p-ATTGTACGTACAAT) in 20 mM TrisHCl, pH 7.5, 5 mM MgCl2, 150 mM NaCl and 2 mM DTT were collected on the EMBL P12 beam line at PETRA III (DESY, Hamburg, Germany) using a Pilatus 6M detector at a sample-detector distance of 3 m and at a wavelength of λ = 0.123981 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). In-line size-exclusion chromatography (SEC) SAS was employed. The SEC parameters were as follows: A 100 μl sample at 8.9 mg/ml (175 μM of complex) was injected at a 0.50 ml/min flow rate onto a GE Superdex 200 Increase 10/300 column at 20°C. 24 successive 0.995 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.

Out of 3000 frames collected during the SEC- run, the sample peak frames corresponded to 1658-1611; buffer frames 2828-2929.

Piwi protein AF_1318 (AfAgo)
Mol. type   Protein
Organism   Archaeoglobus fulgidus
Olig. state   Dimer
Mon. MW   50.9 kDa
 
UniProt   O28951
Sequence   FASTA
 
5'-phosphorilated 14-mer DNA oligoduplex (MZ1289)
Mol. type   DNA
Olig. state   Dimer
Mon. MW   4.3 kDa
Sequence   FASTA
 
PDB ID   1YTU
 
PDB ID   2W42