A mutation affecting a second component of the F0 portion of the magnesium ion-stimulated adenosine triphosphatase of Escherichia coli K12. The uncC424 allele

A new mutant strain of Escherichia coli in which phosphorylation is uncoupled from electron transport was isolated. The new mutant strain has a similar phenotype to the uncB mutant described previously; results from reconstitution experiments in vitro indicate that the new mutation also affects a component of the F0 portion of the Mg2+-stimulated adenosine triphosphatase. A method was developed to incorporate mutant unc alleles into plasmids. Partial diploid strains were prepared in which the uncB402 allele was incorporated into the plasmid and the new unc mutation into the chromosome, or vice versa. Complementation between the mutant unc alleles was indicated by growth on succinate, growth yields on glucose, ATP-dependent transhydrogenase activities, ATP-induced atebrin-fluorescence quenching and oxidative-phosphorylation measurements. The gene in which the new mutation occurs is therefore distinct from the uncB gene, and the mutant allele was designated uncC424.

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Genetic complementation between two mutant unc alleles (unc A401 and unc D409) affecting the F1 portion of the magnesium ion-stimulated adenosine triphosphatase of Escherichia coli K12

Biochemical Journal ◽

10.1042/bj1700593 ◽

1978 ◽

Vol 170 (3) ◽

pp. 593-598 ◽

Cited By ~ 22

Author(s):

G B Cox ◽

J A Downie ◽

F Gibson ◽

J Radik

Keyword(s):

Escherichia Coli ◽

Electron Transport ◽

Fluorescence Quenching ◽

Oxidative Phosphorylation ◽

Mutant Allele ◽

Adenosine Triphosphatase ◽

The Other ◽

Genetic Complementation ◽

Magnesium Ion ◽

Adenosine Triphosphatase Activity

A new mutant strain of Escherichia coli in which phosphorylation is uncoupled from electron transport was isolated. A genetic-complementation analysis, using partial diploid strains, showed that the new mutant allele, uncD409, is in a gene distinct from the other previously identified genes uncA, uncB and uncC. A strain carrying the uncd409 allele has no Mg2+ ion-stimulated adenosine triphosphatase activity and is therefore phenotypically similar to strains carrying the uncA401 mutant allele. Complementation between the uncA401 and the uncD409 alleles occurred, as indicated by growth of partial diploid strains on succinate and their growth yields on limiting concentrations of glucose. Complementation was confirmed by using membranes prepared from the above partial diploids. Such membranes were found to have Mg2+-stimulated adenosine triphosphatase activity, ATP-dependent transhydrogenase activity ADP-induced atebrin-fluorescence quenching and low but significant amounts of oxidative phosphorylation.

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Reconstitution of the energy-linked transhydrogenase activity in membranes from a mutant strain of Escherichia coli K12 lacking magnesium ion- or calcium ion-stimulated adenosine triphosphatase

Biochemical Journal ◽

10.1042/bj1320689 ◽

1973 ◽

Vol 132 (4) ◽

pp. 689-695 ◽

Cited By ~ 23

Author(s):

G. B. Cox ◽

F. Gibson ◽

L. M. McCann ◽

J. D. Butlin ◽

F. L. Crane

Keyword(s):

Escherichia Coli ◽

Mutant Strain ◽

Calcium Ion ◽

Mutant Allele ◽

Adenosine Triphosphatase ◽

Gel Filtration ◽

Magnesium Ion ◽

Adenosine Triphosphatase Activity ◽

Escherichia Coli K12

1. We have isolated a mutant of Escherichia coli K12 (strain AN295) that forms de-repressed amounts of Mg2+,Ca2+-stimulated adenosine triphosphatase. 2. The Mg2+,Ca2+-stimulated triphosphatase activity was separated from membrane preparations from strain AN295 by extraction with 5mm-Tris–HCl buffer containing EDTA and dithiothreitol, resulting in a loss of the ATP-dependent transhydrogenase activity. The non-energy-linked transhydrogenase activity remained in the membrane residue. 3. The solubilized Mg2+,Ca2+-stimulated adenosine triphosphatase activity from strain AN295 was partially purified by repeated gel filtration. The addition of the purified Mg2+,Ca2+-stimulated adenosine triphosphatase to the membrane residue from strain AN295 reactivated the ATP-dependent transhydrogenase activity. 4. Strain AN296, lacking Mg2+,Ca2+-stimulated adenosine triphosphatase activity, was derived by transducing the mutant allele, uncA401, into strain AN295. The ATP-dependent transhydrogenase activity was lost but the non-energy linked transhydrogenase was retained. 5. The ATP-dependent transhydrogenase activity in membrane preparations from strain AN296 (uncA-) could not be re-activated by the purified Mg2+,Ca2+-stimulated adenosine triphosphatase from strain AN295. However, after extraction by 5mm-Tris–HCl buffer containing EDTA and dithiothreitol, the ATP-dependent transhydrogenase activity could be re-activated by the addition of the purified Mg2+,Ca2+-stimulated adenosine triphosphatase from strain AN295 to the membrane residue from strain AN296 (uncA-).

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

Biochemical Journal ◽

10.1042/bj1380211 ◽

1974 ◽

Vol 138 (2) ◽

pp. 211-215 ◽

Cited By ~ 14

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.

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Differential Suppression of priA2::kan Phenotypes in Escherichia coli K-12 by Mutations in priA, lexA, and dnaC

Genetics ◽

10.1093/genetics/143.1.5 ◽

1996 ◽

Vol 143 (1) ◽

pp. 5-13 ◽

Cited By ~ 21

Author(s):

Steven J Sandler ◽

Hardeep S Samra ◽

Alvin J Clark

Keyword(s):

Escherichia Coli ◽

Dna Replication ◽

Mutant Allele ◽

Wild Type ◽

Suppressor Mutations ◽

Dnab Helicase ◽

K 12 ◽

Extragenic Suppressors ◽

Assembly Activity

Abstract First identified as an essential component of the ϕX174 in vitro DNA replication system, PriA has ATPase, helicase, translocase, and primosome-assembly activities. priA1::kan strains of Escherichia coli are sensitive to UV irradiation, deficient in homologous recombination following transduction, and filamentous. priA2::kan strains have eightfold higher levels of uninduced SOS expression than wild type. We show that (1) priA1::kan strains have eightfold higher levels of uninduced SOS expression, (2) priA2::kan strains are UVS and Rec−, (3) lexA3 suppresses the high basal levels of SOS expression of a priA2::kan strain, and (4) plasmid-encoded priA300 (K230R), a mutant allele retaining only the primosome-assembly activity of priA+, restores both UVR and Rec+ phenotypes to a priA2::kan strain. Finally, we have isolated 17 independent UVR Rec+ revertants of priA2::kan strains that carry extragenic suppressors. All 17 map in the C-terminal half of the dnaC gene. DnaC loads the DnaB helicase onto DNA as a prelude for primosome assembly and DNA replication. We conclude that priA's primosome-assembly activity is essential for DNA repair and recombination and that the dnaC suppressor mutations allow these processes to occur in the absence of priA.

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Partial diploids of Escherichia coli carrying normal and mutant alleles affecting oxidative phosphorylation

Biochemical Journal ◽

10.1042/bj1620665 ◽

1977 ◽

Vol 162 (3) ◽

pp. 665-670 ◽

Cited By ~ 46

Author(s):

F Gibson ◽

G B Cox ◽

J A Downie ◽

J Radik

Keyword(s):

Escherichia Coli ◽

Fluorescence Quenching ◽

Oxidative Phosphorylation ◽

Atpase Activity ◽

Adenosine Triphosphatase ◽

Growth Yields ◽

Normal Allele ◽

Adenosine Triphosphatase Activity

A plasmid was isolated which included the region of the Escherichia coli chromosome carrying the known genes concerned with oxidative phosphorylation (unc genes). This plasmid was used to prepare partial diploids carrying normal unc alleles on the episome and one of the three mutant alleles (unc A401, uncB402 or unc-405) on the chromosome. These strains were compared with segregants from which the plasmid had been lost. Dominance of either normal ormutant unc alleles was determined by growth on succinate, growth yields on glucose, Mg-ATPase (Mg2+-stimulated adenosine triphosphatase) activity, atebrin-fluorescence quenching, ATP-dependent transhydrogenase activity and oxidative phosphorylation. In all the above tests, dominance of the normal allele was observed. However, in membranes from the diploid strains which carried a normal allele and either of the mutant alleles affecting Mg-ATPase activity (uncA401 or unc-405), the energy-linked functions were only partially restored.

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A Mutant Allele of rpoD Results in Increased Conversion of Aminoimidazole Ribotide to Hydroxymethyl Pyrimidine in Salmonella enterica

Journal of Bacteriology ◽

10.1128/jb.186.12.4034-4037.2004 ◽

2004 ◽

Vol 186 (12) ◽

pp. 4034-4037 ◽

Cited By ~ 8

Author(s):

Michael J. Dougherty ◽

Diana M. Downs

Keyword(s):

Escherichia Coli ◽

Salmonella Enterica ◽

Mutant Allele ◽

Biosynthetic Pathway ◽

Metabolic Phenotype ◽

Mutant Protein ◽

Cellular Processes ◽

Pyrimidine Moiety

ABSTRACT An allele of rpoD (rpoD1181) that results in increased synthesis of the pyrimidine moiety of thiamine in Salmonella enterica was identified. The S508Y substitution caused by rpoD1181 is analogous to the S506F derivative of the Escherichia coli protein. The properties of this E. coli mutant protein have been well characterized in vitro. Identification of a metabolic phenotype caused by the rpoD1181 allele of S. enterica allows past in vitro results to be incorporated in continuing efforts to understand cellular processes that are integrated with the thiamine biosynthetic pathway.

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Efficacy of trimethoprim in murine experimental infection with a thymidine kinase-deficient mutant of Escherichia coli.

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.41.5.1042 ◽

1997 ◽

Vol 41 (5) ◽

pp. 1042-1045 ◽

Cited By ~ 6

Author(s):

T Tokunaga ◽

K Oka ◽

A Takemoto ◽

Y Ohtsubo ◽

N Gotoh ◽

...

Keyword(s):

Escherichia Coli ◽

Antimicrobial Activity ◽

Thymidine Kinase ◽

Mutant Strain ◽

Experimental Infection ◽

Therapeutic Efficacy ◽

Parent Strain ◽

E Coli

The antimicrobial activity of trimethoprim is antagonized by thymidine in in vitro susceptibility tests. The purpose of this investigation was to determine whether this antagonism also occurred during experimental infection in mice, which have high serum thymidine concentrations. We derived a mutant strain of Escherichia coli, TT-48, incapable of utilizing exogenous thymidine from parent strain E. coli KC-14 and then investigated the in vitro and in vivo antimicrobial activities of trimethoprim, sulfamethoxazole, cefdinir, and ofloxacin against these strains. E. coli TT-48 lacked the activity of thymidine kinase, which catalyzes the conversion of thymidine to thymidylate, but its growth curve remained close to that of the parent strain. The MICs of all of the antimicrobial agents tested, except cefdinir, for the mutant strain were slightly inferior to those for the parent strain. The bactericidal effect of trimethoprim against the parent strain was antagonized by thymidine at concentrations of more than 1 microg/ml, while that against the mutant strain was not affected by thymidine even at the highest concentration (10 microg/ml). The therapeutic efficacy of trimethoprim in experimental murine infections was significantly higher when the mutant rather than the parent strain was used, whereas the therapeutic efficacy of cefdinir or ofloxacin, whose antimicrobial action is independent of folic acid synthesis, was the same with both strains. Unexpectedly, sulfamethoxazole also had similar efficacy against both strains. Thus, high thymidine concentrations antagonized the antimicrobial activity of trimethoprim in vitro and in vivo.

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Positive control of sulphate reduction in Escherichia coli. The nature of the pleiotropic cysteineless mutants of E. coli K 12

Biochemical Journal ◽

10.1042/bj1100597 ◽

1968 ◽

Vol 110 (3) ◽

pp. 597-602 ◽

Cited By ~ 21

Author(s):

M. C. Jones-Mortimer

Keyword(s):

Escherichia Coli ◽

Mutant Allele ◽

Positive Control ◽

Wild Type Allele ◽

Wild Type ◽

E Coli ◽

Type Allele ◽

K 12 ◽

Sulphite Reductase

1. The function of the wild-type alleles of the pleiotropic mutants cysB and cysE of Escherichia coli was investigated. 2. The wild-type allele cysB+ is dominant to the mutant allele cysB in stable and transient heterozygotes. 3. The wild-type allele cysE+ is dominant to the mutant allele cysE, as predicted. 4. Sulphur-starved cultures of cysB or cysE strains contain less than 0·2nmole of free cysteine/mg. dry wt. 5. Complementation in vitro is not observed between extracts of cysB mutants and mutants lacking sulphite reductase only. 6. A scheme, involving positive control of the enzymes of sulphate activation and reduction, is suggested to account for the control of cysteine biosynthesis.

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