Decoupling of size and shape fluctuations in heteropolymeric sequences reconciles discrepancies in SAXS vs. FRET measurements.

Fuertes G Banterle N, Ruff KM, Chowdhury A, Mercadante D, Koehler C, Kachala M, Estrada Girona G, Milles S, Mishra A, Onck PR, Gräter F, Esteban-Martín S, Pappu RV, Svergun DI, Lemke EA, Proc Natl Acad Sci U S A 114(31):E6342-E6351 (2017) Europe PMC

SASDEH2 – Unlabeled nucleoporin NUP49/NSP49 (N49) without denaturant

Nucleoporin NUP49/NSP49
MWexperimental 4 kDa
MWexpected 4 kDa
VPorod 4 nm3
log I(s) 7.61×101 7.61×100 7.61×10-1 7.61×10-2
Nucleoporin NUP49/NSP49 small angle scattering data  s, nm-1
ln I(s)
Nucleoporin NUP49/NSP49 Guinier plot ln 7.61×101 Rg: 1.6 nm 0 (1.6 nm)-2 s2
(sRg)2I(s)/I(0)
Nucleoporin NUP49/NSP49 Kratky plot 1.104 0 3 sRg
p(r)
Nucleoporin NUP49/NSP49 pair distance distribution function Rg: 1.7 nm 0 Dmax: 6 nm

Data validation


Fits and models


log I(s)
 s, nm-1
Unlabeled nucleoporin NUP49/NSP49 (N49) without denaturant Rg histogram Rg, nm
Nucleoporin NUP49/NSP49 OTHER model
Nucleoporin NUP49/NSP49 OTHER model
Nucleoporin NUP49/NSP49 OTHER model
Nucleoporin NUP49/NSP49 OTHER model
Nucleoporin NUP49/NSP49 OTHER model
Nucleoporin NUP49/NSP49 OTHER model
Nucleoporin NUP49/NSP49 OTHER model
Nucleoporin NUP49/NSP49 OTHER model
Nucleoporin NUP49/NSP49 OTHER model

Synchrotron SAXS data from solutions of Unlabeled nucleoporin NUP49/NSP49 (N49) without denaturant in PBS, 10 mM DTT, pH 7.4 were collected on the EMBL P12 beam line at the PETRA III storage ring (DESY; Hamburg, Germany) using a Pilatus 2M detector at a sample-detector distance of 3 m and at a wavelength of λ = 0.1 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). Solute concentrations ranging between 2 and 10 mg/ml were measured at 23°C. 20 successive 0.050 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 low angle data collected at lower concentration were merged with the highest concentration high angle data to yield the final composite scattering curve.

The protein contains a penultimate non-canonical amino acid p-acetylphenylalanine (207 Da) that is represented as U (selenocysteine, 168 Da) in the amino acid sequence for the entry. Therefore, the calculated MW from sequence (MW(expected)) must be adjusted accordingly (ca. 40 Da).

Tags: idp
Nucleoporin NUP49/NSP49 (N49)
Mol. type   Protein
Organism   Saccharomyces cerevisiae
Olig. state   Monomer
Mon. MW   3.9 kDa
 
UniProt   Q02199 (121-154)
Sequence   FASTA