Conformational analysis of a genetically encoded FRET biosensor by SAXS.

Mertens HD, Piljić A, Schultz C, Svergun DI, Biophys J 102(12):2866-75 (2012) Europe PMC


CYNEX4 FRET probe, (eYFP-AnnexinA4-eCFP)
MWI(0) 84 kDa
MWexpected 93 kDa
VPorod 135 nm3
log I(s) 8.69×101 8.69×100 8.69×10-1 8.69×10-2
CYNEX4 FRET probe, (eYFP-AnnexinA4-eCFP) small angle scattering data  s, nm-1
ln I(s)
CYNEX4 FRET probe, (eYFP-AnnexinA4-eCFP) Guinier plot ln 8.69×101 Rg: 3.7 nm 0 (3.7 nm)-2 s2
CYNEX4 FRET probe, (eYFP-AnnexinA4-eCFP) Kratky plot 1.104 0 3 sRg
CYNEX4 FRET probe, (eYFP-AnnexinA4-eCFP) pair distance distribution function Rg: 3.8 nm 0 Dmax: 12.8 nm

Data validation

Fits and models

log I(s)
 s, nm-1
CYNEX4 FRET probe, (eYFP-AnnexinA4-eCFP) DAMMIF model

log I(s)
 s, nm-1
CYNEX4 FRET probe, (eYFP-AnnexinA4-eCFP) CORAL model

Synchrotron SAXS data from solutions of CYNEX4 in 50 mM HEPES 150 mM NaCl 1 mM EGTA, pH 7.5 were collected on the EMBL X33 beam line at the DORIS III storage ring (Hamburg, Germany) using a Pilatus 1M-W detector at a sample-detector distance of 2.7 m and at a wavelength of λ = 0.15 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). Solute concentrations ranging between 0.8 and 12.8 mg/ml were measured at 10°C. Eight successive 15 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.

CYNEX4 FRET probe, (eYFP-AnnexinA4-eCFP) (CYNEX4)
Mol. type   Protein
Organism   Homo sapiens
Olig. state   Monomer
Mon. MW   92.8 kDa
Sequence   FASTA