scholarly journals Properties of membranes from mutant strains of Escherichia coli in which the β-subunit of the adenosine triphosphatase is abnormal

1979 ◽  
Vol 180 (1) ◽  
pp. 111-118 ◽  
Author(s):  
A E Senior ◽  
D R Fayle ◽  
J A Downie ◽  
F Gibson ◽  
G B Cox
Keyword(s):  
Escherichia Coli ◽  
Atpase Activity ◽  
Mutant Allele ◽  
Adenosine Triphosphatase ◽  
Beta Subunit ◽  
Beta Subunits ◽  
F1 Atpase ◽  
Mutant Strains ◽  
Membrane Bound ◽  
Phenotypic Variations

Five uncoupled mutant strains of Escherichia coli carrying mutations in the uncD gene have been studied. In each of these mutant strains the beta-subunit of the F1 portion of the membrane-bound adenosine triphosphatase is abnormal. In one of the mutant strains (carrying the uncD12 allele) in F1-ATPase aggregate was formed which was purified and found to have low ATPase activity. ATPase activity was absent in the other four strains and the abnormal beta-subunits were tightly bound to the membranes. However, membranes from these strains exhibited various proton permeabilities as indicated by NADH-dependent atebrin-fluorescence quenching and bound different amounts of normal F1-ATPase. The amounts of reconstitution of energy-linked reactions after the addition of normal F1-ATPase also varied depending on the mutant allele. It is apparent that considerable phenotypic variations can occur between strains carrying mutations in the same unc gene.

scholarly journals Oxidative phosphorylation in Escherichia coli. Characterization of mutant strains in which F1-ATPase contains abnormal β-subunits

1983 ◽  
Vol 210 (2) ◽  
pp. 395-403 ◽  
Author(s):  
A E Senior ◽  
L Langman ◽  
G B Cox ◽  
F Gibson
Keyword(s):  
Escherichia Coli ◽  
Ionic Strength ◽  
Atpase Activity ◽  
Atp Synthesis ◽  
Beta Subunit ◽  
F1 Atpase ◽  
Dependent Atpase ◽  
Mutant Strains ◽  
Synthesis Activity ◽  
Low Ionic Strength

To facilitate study of the role of the beta-subunit in the membrane-bound proton-translocating ATPase of Escherichia coli, we identified mutant strains from which an F1-ATPase containing abnormal beta-subunits can be purified. Seventeen strains of E. coli, characterized by genetic complementation tests as carrying mutations in the uncD gene (which codes for the beta-subunit), were studied. The majority of these strains (11) were judged to be not useful, as their membranes lacked ATPase activity, and were either proton-permeable as prepared or remained proton-impermeable after washing with buffer of low ionic strength. A further two strains were of a type not hitherto reported, in that their membranes had ATPase activity, were proton-impermeable as prepared, and were not rendered proton-permeable by washing in buffer of low ionic strength. Presumably in these two strains F1-ATPase is not released in soluble form by this procedure. F1-ATPase of normal molecular size were purified from strains AN1340 (uncD478), AN937 (uncD430), AN938 (uncD431) and AN1543 (uncD484). F1-ATPase from strain AN1340 (uncD478) had 15% of normal specific Mg-dependent ATPase activity and 22% of normal ATP-synthesis activity. The F1-ATPase preparations from strains AN937, AN938 and AN1543 had respectively 1.7%, 1.8% and 0.2% of normal specific Mg-dependent ATPase activity, and each of these preparations had very low ATP-synthesis activity. The yield of F1-ATPase from the four strains described was almost twice that obtained from a normal haploid strain. The kinetics of Ca-dependent ATPase activity were unusual in each of the four F1-ATPase preparations. It is likely that these four mutant uncD F1-ATPase preparations will prove valuable for further experimental study of the F1-ATPase catalytic mechanism.

scholarly journals Oxidative phosphorylation in Escherichia coli K12. An uncoupled mutant with altered membrane structure

1974 ◽  
Vol 138 (2) ◽  
pp. 211-215 ◽  
Author(s):  
G. B. Cox ◽  
F. Gibson ◽  
L. McCann
Keyword(s):  
Escherichia Coli ◽  
Electron Transport ◽  
Membrane Structure ◽  
Mutant Allele ◽  
Adenosine Triphosphatase ◽  
Mineral Salts ◽  
Normal Cells ◽  
E Coli ◽  
Escherichia Coli K12 ◽  
Membrane Bound

1. A new mutant strain (AN228) of Escherichia coli K12, unable to couple phosphorylation to electron transport, has been isolated. The mutant allele (unc-405), in strain AN228, was found to map near the uncA and uncB genes at about minute 74 on the E. coli genome. 2. A transductant strain (AN285) carrying the unc-405 allele is similar to the uncA and uncB mutants described previously in that it is unable to grow on succinate, gives a low aerobic yield on limiting concentrations of glucose, has a normal rate of electron transport, is unable to couple phosphorylation to electron transport, and lacks ATP-dependent transhydrogenase activity. 3. Strain AN285 (unc-405) is similar to an uncA mutant, but different from an uncB mutant, in that it is unable to grow anaerobically in a glucose–mineral-salts medium, and membrane preparations do not have Mg2+-stimulated adenosine triphosphatase activity. 4. Strain AN285 (unc-405) does not form an aggregate analogous to the membrane-bound Mg2+-stimulated adenosine triphosphatase aggregate found in normal cells. In this respect it differs from strain AN249 (uncA−), which forms an inactive membrane-bound Mg2+-stimulated adenosine triphosphatase aggregate.

scholarly journals Characterization of the mutant-unc D-gene product in a strain of Escherichia coli K12. An altered β-subunit of the magnesium ion-stimulated adenosine triphosphatase

1978 ◽  
Vol 172 (3) ◽  
pp. 523-531 ◽  
Author(s):  
D R H Fayle ◽  
J A Downie ◽  
G B Cox ◽  
F Gibson ◽  
J Radik
Keyword(s):  
Escherichia Coli ◽  
Adenosine Triphosphatase ◽  
Polyacrylamide Gel Electrophoresis ◽  
Beta Subunit ◽  
Beta Subunits ◽  
Normal Strain ◽  
Diploid Strain ◽  
Escherichia Coli K12 ◽  
Low Ionic Strength

Membranes from a mutant strain of Escherichia coli K12 carrying the uncD409 allele were washed in low-ionic-strength buffers in the presence or absence of the proteinase inhibitor p-aminobenzamidine. Unlike membranes from a normal strain, those from strain AN463 (uncD409) did not become proton-permeable, as judged by NADH-induced atebrinfluorescence quenching, when the membranes were washed in the absence of p-aminobenzamide. Furthermore, ATP-dependent atebrin-fluorscence quenching in such washed membranes could not be reconstituted by the addition of solubilized Mg2+-stimulated adenosine triphosphatase preparations. The examination by two-dimensional polyacrylamide-gel electrophoresis of the polypeptide composition of the washed membranes from strain AN463 (uncD409) indicated the presence of a polypeptide of similar molecular weight to the normal beta-subunit of the Mg2+-stimulated adenosine triphosphatase, but with an altered isoelectric point. Both the normal and abnormal beta-subunits were identified in membranes prepared from a partial diploid strain carrying both the unc+ and uncD409 alleles. It is concluded that the uncD gene codes for the beta-subunit of the Mg2+-stimulated adenosine triphosphatase.

Binding of [14C]aurovertin D to Escherichia coli F1-ATPase and the isolated .beta. subunit. Correlation with inhibition of the ATPase activity

Biochemistry ◽  
1983 ◽  
Vol 22 (14) ◽  
pp. 3485-3492 ◽  
Author(s):  
Jean Paul Issartel ◽  
Gerard Klein ◽  
Michel Satre ◽  
Pierre V. Vignais
Keyword(s):  
Escherichia Coli ◽  
Atpase Activity ◽  
Beta Subunit ◽  
F1 Atpase

scholarly journals The uncA gene codes for the α-subunit of the adenosine triphosphatase of Escherichia coli. Electrophoretic analysis of uncA mutant strains

1979 ◽  
Vol 180 (1) ◽  
pp. 103-109 ◽  
Author(s):  
A E Senior ◽  
J A Downie ◽  
G B Cox ◽  
F Gibson ◽  
L Langman ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Ionic Strength ◽  
Adenosine Triphosphatase ◽  
Alpha Subunit ◽  
Subunit Structure ◽  
Net Charge ◽  
Α Subunit ◽  
F1 Atpase ◽  
Mutant Strains ◽  
Low Ionic Strength

Four mutant strains of Escherichia coli which lack membrane-bound adenosine triphosphatase activity were shown by genetic-complementation tests to carry mutations in the uncA gene. A soluble inactive F1-ATPase aggregate was released from the membranes of three of the uncA mutant strains by low-ionic-strength washing, and purified by procedures developed for the purification of F1-ATPase from normal strains. Analysis of the subunit structure by two-dimensional gel electrophoresis indicated that the F1-ATPase in strains carrying the uncA401 or uncA453 alleles had a subunit structure indistinguishable from normal F1-ATPase. In contrast, the F1-ATPase from the strain carrying the uncA447 allele contained an alpha-subunit of normal molecular weight, but abnormal net charge. Membranes from strains carrying the uncA450 allele did not have F1-ATPase aggregates that could be solubilized by low-ionic-strength washing. However, a partial dipolid strain carrying both the uncA+ and uncA450 alleles formed an active F1-ATPase aggregate which could be solubilized by low-ionic-strength washing of the membranes and which contained two types of alpha-subunit, one of which was normal and the other had abnormal net charge. It is concluded that the uncA gene codes for the alpha-subunit of the adenosine triphosphatase.

scholarly journals An additional acidic residue in the membrane portion of the b-subunit of the energy-transducing adenosine triphosphatase of Escherichia coli affects both assembly and function

1984 ◽  
Vol 221 (1) ◽  
pp. 43-51 ◽  
Author(s):  
D A Jans ◽  
A L Fimmel ◽  
L Hatch ◽  
F Gibson ◽  
G B Cox
Keyword(s):  
Escherichia Coli ◽  
Oxidative Phosphorylation ◽  
Atpase Activity ◽  
Dna Sequence ◽  
Mutant Allele ◽  
Adenosine Triphosphatase ◽  
The Other ◽  
Multicopy Plasmid ◽  
B Subunit ◽  
And Function

Glycine at position 9 is replaced by aspartic acid in the mutant b-subunit of Escherichia coli F1F0-ATPase coded for by the uncF476 allele. The mutant b-subunit is not assembled into the membrane in haploid strains carrying the uncF476 allele, but, if the mutant allele is incorporated into a multicopy plasmid, then some assembly of the mutant b-subunit occurs. Two revertant strains were characterized, one of which (AN2030) was a full revertant, the other (AN1953) a partial revertant. DNA sequencing indicated that in strain AN2030 the uncF476 mutation had reverted to give the sequence found in the normal uncF gene. The partial-revertant strain AN1953, however, retained the DNA sequence of the uncF476 allele, and complementation analysis indicated that the second mutation may be in the uncA gene. Membranes prepared from the partial-revertant strain carried out oxidative phosphorylation, although the membranes appeared to be impermeable to protons, and the ATPase activity was sensitive to the inhibitor dicyclohexylcarbodi-imide.

scholarly journals Lysine 155 in beta-subunit is a catalytic residue of Escherichia coli F1 ATPase.

1993 ◽  
Vol 268 (10) ◽  
pp. 6989-6994
Author(s):  
A.E. Senior ◽  
S. Wilke-Mounts ◽  
M.K. al-Shawi
Keyword(s):  
Escherichia Coli ◽  
Beta Subunit ◽  
Catalytic Residue ◽  
F1 Atpase
Sign in / Sign up

Export Citation Format

Share Document