Rational thermostabilisation of four-helix bundle dimeric de novo proteins.

Irumagawa S, Kobayashi K, Saito Y, Miyata T, Umetsu M, Kameda T, Arai R, Sci Rep 11(1):7526 (2021) Europe PMC

SASDKM8 – Rationally optimised WA20 (ROWA) dimer

Rationally optimised WA20 mutant N22A/H86K (ROWA) dimer
MWI(0) 25 kDa
MWexpected 25 kDa
VPorod 30 nm3
log I(s) 5.90×10-2 5.90×10-3 5.90×10-4 5.90×10-5
Rationally optimised WA20 mutant N22A/H86K (ROWA) dimer small angle scattering data  s, nm-1
ln I(s)
Rationally optimised WA20 mutant N22A/H86K (ROWA) dimer Guinier plot ln 5.90×10-2 Rg: 2.5 nm 0 (2.5 nm)-2 s2
(sRg)2I(s)/I(0)
Rationally optimised WA20 mutant N22A/H86K (ROWA) dimer Kratky plot 1.104 0 3 sRg
p(r)
Rationally optimised WA20 mutant N22A/H86K (ROWA) dimer pair distance distribution function Rg: 2.6 nm 0 Dmax: 9.9 nm

Data validation


Fits and models


log I(s)
 s, nm-1
Rationally optimised WA20 mutant N22A/H86K (ROWA) dimer DAMMIN model

log I(s)
 s, nm-1
Rationally optimised WA20 mutant N22A/H86K (ROWA) dimer GROMACS model

Synchrotron SAXS data from solutions of Rationally optimised WA20 (ROWA) dimer in 20 mM HEPES, 150 mM NaCl, 5% glycerol,, pH 7.5 were collected on the BL-10C beam line at the Photon Factory (PF), High Energy Accelerator Research Organization (KEK) storage ring (Tsukuba, Japan) using a Pilatus3 2M detector at a sample-detector distance of 1.1 m and at a wavelength of λ = 0.13 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). One solute concentration of 3.93 mg/ml was measured at 20°C. 30 successive 5 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.

Rationally optimised WA20 mutant N22A/H86K (ROWA) dimer (ROWA dimer)
Mol. type   Protein
Organism   Artificial protein (de novo protein)
Olig. state   Dimer
Mon. MW   12.5 kDa
Sequence   FASTA
 
PDB code   3VJF