Small-angle X-ray scattering structural insights into alternative pathway of actin oligomerization associated with inactivated state

Ryzhykau Y, Povarova O, Dronova E, Kuklina D, Antifeeva I, Ilyinsky N, Okhrimenko I, Semenov Y, Kuklin A, Ivanovich V, Fonin A, Uversky V, Turoverov K, Kuznetsova I, Biochemical and Biophysical Research Communications :149340 (2023) DOI

SASDTZ4 – I-actin - Skeletal muscle actin from Oryctolagus cuniculus: oligomer of time-inactivated actin (peak SEC fraction)

Actin, alpha skeletal muscle
MWexperimental 355 kDa
MWexpected 42 kDa
VPorod 549 nm3
log I(s) 7.62×10-1 7.62×10-2 7.62×10-3 7.62×10-4
Actin, alpha skeletal muscle small angle scattering data  s, nm-1
ln I(s)
Actin, alpha skeletal muscle Guinier plot ln 7.62×10-1 Rg: 4.9 nm 0 (4.9 nm)-2 s2
(sRg)2I(s)/I(0)
Actin, alpha skeletal muscle Kratky plot 1.104 0 √3 sRg
p(r)
Actin, alpha skeletal muscle pair distance distribution function Rg: 5.7 nm 0 Dmax: 22 nm

Data validation

Fits and models

log I(s)
 s, nm-1
Actin, alpha skeletal muscle OTHER [STATIC IMAGE] model
SAXS data from solutions of I-actin from Oryctolagus cuniculus (an oligomer of time-inactivated actin) in 5 mM Tris/Tris-HCl, 2.0 mM EDTA, pH 8.1 were collected using a Rigaku MicroMax 007-HF instrument at the Moscow Institute of Physics and Technology (MIPT; Dolgoprudny, Russian Federation) equipped with a multiwire gas-filled ASM DTR Triton 200 detector at a sample-detector distance of 2 m and at a wavelength of λ = 0.15406 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). One solute concentration of 0.74 mg/ml was measured at 20°C. One 7200 second frame was collected. The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted.

For I-actin, the model of an elliptical cylinder proved to be applicable. Oligomers of I-actin have length of ~14 nm, that is significantly shorter compared to F-actin. Regarding the cross-sections, the axes of the ellipses were 5.2 nm and 8.5 nm for F-actin, and 5.6 nm and 10.8 nm for I-actin. The cross-sectional area of I-actin is ~40% larger than F-actin. This indicates another structure of oligomers, and, therefore, fundamentally different oligomerization mechanism in I-actin compared to F-actin.

Actin, alpha skeletal muscle
Mol. type   Protein
Organism   Oryctolagus cuniculus
Olig. state   Monomer
Mon. MW   42.1 kDa
 
UniProt   P68135 (1-377)
Sequence   FASTA