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Literature summary for 7.5.2.6 extracted from

  • Schultz, K.; Merten, J.; Klug, C.
    Effects of the L511P and D512G mutations on the Escherichia coli ABC transporter MsbA (2011), Biochemistry, 50, 2594-2602 .
    View publication on PubMedView publication on EuropePMC
Search for more literature for 7.5.2.6

Cloned(Commentary)

Cloned (Comment) Organism
gene msbA, recombinant expression of N-terminally His6-tagged wild-type and mutant enzymes in Escherichia coli NovaBlue cells Escherichia coli

Crystallization (Commentary)

Crystallization (Comment) Organism
apo,MgADP/Vi-bound and MgAMP-PNP-bound MsbA, X-ray diffraction structure determination and analysis Escherichia coli

Protein Variants

Protein Variants Comment Organism
A270T site-directed mutagenesis, the mutation causes the protein to become inactive at high temperatures Escherichia coli
D512G random mutagenesis, the mutant is able to hydrolyze ATP 3fold faster than the wild-type enzyme Escherichia coli
I385C site-directed mutagenesis of the spin-labeled reporter site Escherichia coli
I385C/D512G site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes slight changes upon MgATP/Vi binding Escherichia coli
I385C/L511P site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding Escherichia coli
L504C site-directed mutagenesis of the spin-labeled reporter site Escherichia coli
L504C/D512G site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding, the mutant exhibits a general broadening of the spectrum Escherichia coli
L504C/L511P site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding Escherichia coli
L511P random mutagenesis, the mutant is able to bind ATP at near-wild-type levels but is unable to maintain cell viability in an in vivo growth assay, it is dysfunctional at some point after ATP binding. The L511P mutation prevents effective ATP hydrolysis, only small amounts of ATP are hydrolyzed Escherichia coli
additional information mutant phenotypes, overview Escherichia coli
Q485C site-directed mutagenesis of the spin-labeled reporter site Escherichia coli
Q485C/D512G site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding Escherichia coli
Q485C/L511P site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding Escherichia coli
S380C site-directed mutagenesis of the spin-labeled reporter site Escherichia coli
S380C/D512G site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding and exhibits an additional shift (approximately 30%) toward the immobilized population Escherichia coli
S380C/L511P site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding Escherichia coli
S423C site-directed mutagenesis of the spin-labeled reporter site Escherichia coli
S423C/D512G site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes slight changes upon MgATP/Vi binding Escherichia coli
S423C/L511P site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding Escherichia coli
S482C site-directed mutagenesis of the spin-labeled reporter site Escherichia coli
S482C/D512G site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding Escherichia coli
S482C/L511P site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding Escherichia coli
T541C site-directed mutagenesis of the spin-labeled reporter site Escherichia coli
T541C/D512G site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding Escherichia coli
T541C/L511P site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding Escherichia coli
V426C site-directed mutagenesis of the spin-labeled reporter site Escherichia coli
V426C/D512G site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding Escherichia coli
V426C/L511P site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding, the mutant shows a more immobile spectrum in the presence of MgATP Escherichia coli
V534C site-directed mutagenesis of the spin-labeled reporter site Escherichia coli
V534C/D512G site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes slight changes upon MgATP/Vi binding Escherichia coli
V534C/L511P site-directed mutagenesis, the additional mutation of the spin-labeled reporter site causes changes upon MgATP/Vi binding Escherichia coli

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Escherichia coli

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ATP + H2O + lipid A-core oligosaccharide[side 1] Escherichia coli
-
 ADP + phosphate + lipid A-core oligosaccharide[side 2]
-
 ?
ATP + H2O + lipid A-core oligosaccharide[side 1] Escherichia coli K12
-
 ADP + phosphate + lipid A-core oligosaccharide[side 2]
-
 ?

Organism

Organism UniProt Comment Textmining
Escherichia coli P60752
-
-
Escherichia coli K12 P60752
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant N-terminally His6-tagged wild-type and mutant enzymes from Escherichia coli NovaBlue cells by cobalt affinity chromatography and ultrafiltration Escherichia coli

Renatured (Commentary)

Renatured (Comment) Organism
MsbA is reconstituted into 65:25:10 phosphatidylethanolamine: phosphatidylglycerol: cardiolipin inner membrane liposomes at a 250:1 lipid:protein molar ratio Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP + H2O + lipid A-core oligosaccharide[side 1]
-
 Escherichia coli ADP + phosphate + lipid A-core oligosaccharide[side 2]
-
 ?
ATP + H2O + lipid A-core oligosaccharide[side 1]
-
 Escherichia coli K12 ADP + phosphate + lipid A-core oligosaccharide[side 2]
-
 ?

Subunits

Subunits Comment Organism
homodimer the MsbA homodimer is comprised of two identical monomers, each with a cytosolic nucleotide binding domain (NBD) and a transmembrane domain (TMD) Escherichia coli

Synonyms

Synonyms Comment Organism
MsbA
-
 Escherichia coli

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
 assay at Escherichia coli

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7
-
 assay at Escherichia coli

Cofactor

Cofactor Comment Organism Structure
ATP
-
 Escherichia coli