Direct localization of detergents and bacteriorhodopsin in the lipidic cubic phase by small-angle neutron scattering

Cleveland IV T, Blick E, Krueger S, Leung A, Darwish T, Butler P, IUCrJ 8(1) (2021) DOI

SASDJE4 – Bacteriorhodopsin in contrast-matched lipidic cubic phase formed with deuterated monoolein

MWexperimental 29 kDa
MWexpected 27 kDa
VPorod 49 nm3
log I(s) 2.10×10-1 2.10×10-2 2.10×10-3 2.10×10-4
Bacteriorhodopsin small angle scattering data  s, nm-1
ln I(s)
Bacteriorhodopsin Guinier plot ln 2.10×10-1 Rg: 2.7 nm 0 (2.7 nm)-2 s2
Bacteriorhodopsin Kratky plot 1.104 0 3 sRg
Bacteriorhodopsin pair distance distribution function Rg: 2.2 nm 0 Dmax: 6.2 nm

Data validation

There are no models related to this curve.

Small-angle neutron scattering (SANS) data from Bacteriorhodopsin (bR) after incorporation into lipidic cubic phase formed by deuterated monoolein, measured at 22°C. After mixing protein solution with deuterated monoolein to form the lipidic cubic phase, the final concentration of bR was 5.20 mg/mL. The final volume fractions of bR, octyl glucoside, and deuterated monoolein in the sample were 0.39%, 1.56%, and 27.6%, with the remainder buffer. The sample buffer consisted of 25 mM Sodium Phosphate and 1.35 mM potassium hydroxide in 100% D2O (in an H2O solution with the same buffer concentrations, the pH is 5.6). Data were collected on the NG7 SANS instrument at the NIST Center for Neutron Research (Gaithersburg, MD, USA) using a 640x640 mm 3He position sensitive Ordela 2660N detector at wavelength of λ = 0.6 nm (I(s) vs s, where s = 4πsinθ/λ and 2θ is the scattering angle). The data were normalized to the intensity of the transmitted beam and radially averaged; the constant incoherent scattering background was determined from the scattering at high angle (s > 4.5 nm^(-1)) and was subtracted. Three detector positions (in m) and count times (s) were employed: 1 m (one frame of 300 s total count time); 4 m (one frame of 1800 s total count time); 13 m (one frame of 600 s total count time). Reduced scattering data in its original format, without any subtractions, is also provided in a downloadable archive. This data is in NIST 6-column format and can be displayed in, for example, the software SasView. These data also include an estimate of the Q-dependent instrument resolution, which can be used during model fitting to smear calculated SANS intensity curves. The following additional scattering data are also present in the archive: (1) A concentration series of bR, solubilized in octyl glucoside and incorporated into deuterated monoolein cubic phase. (2) A concentration series of bR, solubilized in partially-deuterated octyl glucoside mixture and incorporated into deuterated monoolein cubic phase. (3) A panel of 6 detergents incorporated into deuterated monoolein cubic phase, with and without 2 M phosphate precipitant. (4) Deuterated monoolein cubic phase without detergent or bR. (5) Non-deuterated monoolein cubic phase without detergent or bR, in 100% D2O buffer (non-matched) and 7% D2O buffer (matched). (6) SAXS curves of the panel of 6 detergents incorporated into deuterated monoolein cubic phase. All samples were in 100% D2O buffer unless specified otherwise. A spreadsheet containing full details of all sample conditions is included in the archive.

Bacteriorhodopsin (bR)
Mol. type   Protein
Organism   Halobacterium salinarum
Olig. state   Monomer
Mon. MW   26.9 kDa
UniProt   P02945 (14-262)
Sequence   FASTA