SASDDE5: NBD-MsbA (apo) (Nucleotide Binding Domain of Lipid A export ATP-b...

Basic information

Entry

Database: SASBDB / ID: SASDDE5

Sample

NBD-MsbA (apo)

• Nucleotide Binding Domain of Lipid A export ATP-binding/permease protein MsbA (protein), NBD-MsbA, Escherichia coli (strain K12)

Function / homology

Function:

MsbA transporter complex / lipopolysaccharide floppase activity / lipid translocation / ABC-type lipid A-core oligosaccharide transporter / lipopolysaccharide transport / ATPase-coupled lipid transmembrane transporter activity / ABC-type xenobiotic transporter activity / lipid transport / ATP-binding cassette (ABC) transporter complex / transmembrane transport / lipid binding / ATP hydrolysis activity / ATP binding / membrane / identical protein binding / plasma membrane

Domain/homology:

Lipid A export ATP-binding/permease protein msbA family profile. / ABC transporter, lipid A-core flippase, MsbA / Type 1 protein exporter / ABC transporter transmembrane region / ABC transporter type 1, transmembrane domain / ABC transporter integral membrane type-1 fused domain profile. / ABC transporter type 1, transmembrane domain superfamily / ABC transporter-like, conserved site / ABC transporters family signature. / ABC transporter / ABC transporter-like, ATP-binding domain / ATP-binding cassette, ABC transporter-type domain profile. / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase

Component:

ATP-dependent lipid A-core flippase

• Biological species: Escherichia coli (strain K12) (bacteria)
• Citation: Journal: Structure / Year: 2018; Title: Conformational States of ABC Transporter MsbA in a Lipid Environment Investigated by Small-Angle Scattering Using Stealth Carrier Nanodiscs.; Authors: Inokentijs Josts / Julius Nitsche / Selma Maric / Haydyn D Mertens / Martine Moulin / Michael Haertlein / Sylvain Prevost / Dmitri I Svergun / Sebastian Busch / V Trevor Forsyth / Henning Tidow / ; Abstract: Structural studies of integral membrane proteins (IMPs) are challenging, as many of them are inactive or insoluble in the absence of a lipid environment. Here, we describe an approach making use of fractionally deuterium labeled "stealth carrier" nanodiscs that are effectively invisible to low-resolution neutron diffraction and enable structural studies of IMPs in a lipidic native-like solution environment. We illustrate the potential of the method in a joint small-angle neutron scattering (SANS) and X-ray scattering (SAXS) study of the ATP-binding cassette (ABC) transporter protein MsbA solubilized in the stealth nanodiscs. The data allow for a direct observation of the signal from the solubilized protein without contribution from the surrounding lipid nanodisc. Not only the overall shape but also differences between conformational states of MsbA can be reliably detected from the scattering data, demonstrating the sensitivity of the approach and its general applicability to structural studies of IMPs.

Contact author

• Haydyn Mertens (EMBL-Hamburg, European Molecular Biology Laboratory (EMBL) - Hamburg outstation, Notkestraße 85, Geb. 25A, 22607 Hamburg, Deutschland, Germany)

Models

Sample

• Sample: Name: NBD-MsbA (apo) / Contrast: 2.842 / Specific vol: 0.738 / Specimen concentration: 18 mg/ml / Concentration method: A280
• Buffer: Name: 30 mM Tris, 150 mM NaCl, 0.5 mM TCEP / pH: 7.5

Entity #1019

Name: NBD-MsbA / Type: protein
Description: Nucleotide Binding Domain of Lipid A export ATP-binding/permease protein MsbA
Formula weight: 27.263 / Num. of mol.: 1 / Source: Escherichia coli (strain K12) / References: UniProt: P60752
Sequence:

RATGDVEFRN VTFTYPGRDV PALRNINLKI PAGKTVALVG RSGSGKSTIA SLITRFYDID EGEILMDGHD LREYTLASLR NQVALVSQNV HLFNDTVANN IAYARTEQYS REQIEEAARM AYAMDFINKM DNGLDTVIGE NGVLLSGGQR QRIAIARALL RDSPILILDE ATSALDTESE RAIQAALDEL QKNRTSLVIA HRLSTIEKAD EIVVVEDGVI VERGTHNDLL EHRGVYAQLH KMQFGQ

Experimental information

• Beam: Instrument name: PETRA III EMBL P12 / City: Hamburg / 国: Germany / Type of source: X-ray synchrotronSynchrotron / Wavelength: 0.124 Å / Dist. spec. to detc.: 3.3 mm
• Detector: Name: Pilatus 2M

Scan

Title: NBD-MsbA (apo) / Measurement date: May 30, 2017 / Storage temperature: 20 °C / Cell temperature: 20 °C / Exposure time: 0.05 sec. / Number of frames: 20 / Unit: 1/nm /

	Min	Max
Q	0.0611	3.3196

Distance distribution function P(R)

Sofotware P(R): GNOM 5.0 / Number of points: 1175 /

	Min	Max
Q	0.0913838	3.3196
P(R) point	1	1175
R	0	7.3

Result

Type of curve: single_conc / Standard: water

	Experimental	Experimental error	Porod	Porod error
MW	30.12 kDa	5	29.6 kDa	29.6
Volume	-	-	47.39 nm3	5

	P(R)	P(R) error	Guinier	Guinier error
Forward scattering, I0	0.022	0.001	0.022	0.001
Radius of gyration, Rg	2.17 nm	0.1	2.15 nm	0.1

	Min	Max	Error
D	-	7.3	0.5
Guinier point	12	198	-