The missing part: the Archaeoglobus fulgidus Argonaute forms a functional heterodimer with an N-L1-L2 domain protein.

Manakova E Golovinas E, Pocevičiūtė R, Sasnauskas G, Silanskas A, Rutkauskas D, Jankunec M, Zagorskaitė E, Jurgelaitis E, Grybauskas A, Venclovas Č, Zaremba M, Nucleic Acids Res (2024) Europe PMC

SASDRY8 – Single-chain Archaeoglobus fulgidus argonaute fAfAgo-fusion variant complexed with a 14 base pair DNA oligoduplex in low salt buffer

single-chain full Archaeoglobus fulgidus Argonaute
5'-end phosphorylated DNA oligoduplex, 14 bp (MZ1288)
MWI(0) 84 kDa
MWexpected 87 kDa
VPorod 131 nm3
log I(s) 6.72×10-2 6.72×10-3 6.72×10-4 6.72×10-5
single-chain full Archaeoglobus fulgidus Argonaute 5'-end phosphorylated DNA oligoduplex, 14 bp (MZ1288) small angle scattering data  s, nm-1
ln I(s)
single-chain full Archaeoglobus fulgidus Argonaute 5'-end phosphorylated DNA oligoduplex, 14 bp (MZ1288) Guinier plot ln 6.72×10-2 Rg: 3.0 nm 0 (3.0 nm)-2 s2
(sRg)2I(s)/I(0)
single-chain full Archaeoglobus fulgidus Argonaute 5'-end phosphorylated DNA oligoduplex, 14 bp (MZ1288) Kratky plot 1.104 0 3 sRg
p(r)
single-chain full Archaeoglobus fulgidus Argonaute 5'-end phosphorylated DNA oligoduplex, 14 bp (MZ1288) pair distance distribution function Rg: 3.0 nm 0 Dmax: 10.3 nm

Data validation


Fits and models


log I(s)
 s, nm-1
single-chain full Archaeoglobus fulgidus Argonaute 5'-end phosphorylated DNA oligoduplex, 14 bp (MZ1288) GASBOR model

log I(s)
 s, nm-1
single-chain full Archaeoglobus fulgidus Argonaute 5'-end phosphorylated DNA oligoduplex, 14 bp (MZ1288) CUSTOM IN-HOUSE model

log I(s)
 s, nm-1
single-chain full Archaeoglobus fulgidus Argonaute 5'-end phosphorylated DNA oligoduplex, 14 bp (MZ1288) CUSTOM IN-HOUSE model

Synchrotron SAXS data from solutions of a fAfAgo fusion protein complexed with a 14 base pair DNA oligoduplex in 20 mM TrisHCl pH7.5, 200 mM NaCl, 5 mM MgCl2, 1 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 = 0.124 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). One solute concentration of 1.65 mg/ml was measured at 20°C. 33 successive 0.195 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.

DNA oligoduplex was mixed with scfAfAgo in the presence of MgCl2, buffer was changed using a NAP Illustra column and concentrated to 1.65 mg/ml. Note: The DNA sequence of the model shown in this entry is different to the DNA sequence of the oligoduplex in the sample used for SAXS.

single-chain full Archaeoglobus fulgidus Argonaute (sgfAfAgo)
Mol. type   Protein
Organism   Archaeoglobus fulgidus
Olig. state   Monomer
Mon. MW   77.9 kDa
Sequence   FASTA
 
5'-end phosphorylated DNA oligoduplex, 14 bp (MZ1288) (MZ1288)
Mol. type   DNA
Olig. state   Monomer
Mon. MW   8.7 kDa
Sequence   FASTA