Elucidation of the lesions present in the transcription defectivefitA76 mutant ofEscherichia coli: Implication of phenylalanyl tRNA synthetase subunits as transcription factors

Ramalingam, Sandhya; Munavar, M. Hussain; Selvaraj, S.; Ruckmani, A.; Jayaraman, R.

doi:10.1007/bf02941195

Cited by 6 publications

(22 citation statements)

References 23 publications

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“…Surprisingly, the sequence analyses of pheS region in fitA76 revealed the presence of very same G 293 →A 293 mutation leading to glycine to aspartic acid as was reported to be present in the classical translation defective pheS5 Ts mutant. This was totally unexpected and then it was found that not even a single transcriptional abnormality characteristic of fitA76 mutant was true of classical pheS5 Ts mutant . In fact much earlier itself Jabbar and Jayaraman predicted that fitA76 and pheS5 differ in their characteristic phenotype.…”

Section: Discussionmentioning

confidence: 99%

“…However, despite extensive genetic analysis whether fit mutations per se , defined alleles of hitherto unreported genes or defined alleles of other genes mapping in that region but whose involvement in transcription was not known remained unequivocal. Later, Ramalingam et al cloned a 2.1 Kb fragment from fit + E. coli which complemented the Ts phenotype of only fitA and not fitB mutants. Restriction mapping, identification of the protein product encoded by the 2.1 Kb insert together with sequence analyses of the 2.1 Kb fragment, clearly revealed the presence of pheS gene coding for α‐subunit of PheRS in this fragment .…”

Section: Discussionmentioning

confidence: 99%

“…Later, Ramalingam et al cloned a 2.1 Kb fragment from fit + E. coli which complemented the Ts phenotype of only fitA and not fitB mutants. Restriction mapping, identification of the protein product encoded by the 2.1 Kb insert together with sequence analyses of the 2.1 Kb fragment, clearly revealed the presence of pheS gene coding for α‐subunit of PheRS in this fragment . Surprisingly, the sequence analyses of pheS region in fitA76 revealed the presence of very same G 293 →A 293 mutation leading to glycine to aspartic acid as was reported to be present in the classical translation defective pheS5 Ts mutant.…”

Section: Discussionmentioning

confidence: 99%

“…In fact much earlier itself Jabbar and Jayaraman predicted that fitA76 and pheS5 differ in their characteristic phenotype. The paradox was resolved when the fitA76 was found to be a double mutant one is G 293 →A 293 as the case with pheS5 and another mutation named as fit95 expected to be located in pheT . This mutation fit95 , per se confers the Ts phenotype on media devoid of salt.…”

Section: Discussionmentioning

confidence: 99%

“…This mutation fit95 , per se confers the Ts phenotype on media devoid of salt. Taken together, it has been postulated that FitA

\equiv

PheS and FitB

\equiv

PheT and it is both subunits of PheRS enzyme that might participate in transcription control as accessory transcription factors also, a notion yet to be proved by in vitro experimentation .…”

Section: Discussionmentioning

confidence: 99%

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Glu₅₇₁ of PheT plays a pivotal role in the thermal stability of Escherichia coli PheRS enzyme

et al. 2018

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As of date the two temperature sensitive mutations isolated in pheST operon include pheS5 (G →A ) and pheT354. Recently, we reported that G of pheS defines a hot spot for intragenic suppressors of pheS5. In this investigation, in 13 independent experiments, a collection of temperature sensitive mutants were isolated by localized mutagenesis. Complementation using clones bearing pheS , pheT , and pheS T indicated that 34 mutants could harbor lesion(s) in pheS and four could be in pheT and one mutant might be a double mutant. Surprisingly, all the 34 pheS mutants harbored the very same (G →A ) transition mutation as present in the classical pheS5 mutant. Most unexpectedly, the four pheT mutants isolated harbored the same G →A transition, a mutation which is hitherto unreported. Since all the four pheT mutants were defined by the same G →A base change, we believe that getting other mutations could be hard hitting and therefore it is proposed that G itself could be a "hot spot" for emergence of Ts mutations in pheT and similarly G itself could be a "hot spot" for Ts lesions in pheS. These results clearly imply a vital role for Glutamic acid (Glu ) of PheT and reinforce criticality of Glycine (Gly ) of PheS in the thermal stability of PheRS enzyme.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

“…This mutation fit95 , per se confers the Ts phenotype on media devoid of salt. Taken together, it has been postulated that FitA

\equiv

PheS and FitB

\equiv

PheT and it is both subunits of PheRS enzyme that might participate in transcription control as accessory transcription factors also, a notion yet to be proved by in vitro experimentation .…”

Section: Discussionmentioning

confidence: 99%

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Glu₅₇₁ of PheT plays a pivotal role in the thermal stability of Escherichia coli PheRS enzyme

et al. 2018

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G₆₇₃ could be a novel mutational hot spot for intragenic suppressors of pheS5 lesion in Escherichia coli

Ponmani

Munavar

2014

MicrobiologyOpen

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The pheS5 Ts mutant of Escherichia coli defined by a G293 → A293 transition, which is responsible for thermosensitive Phenylalanyl-tRNA synthetase has been well studied at both biochemical and molecular level but genetic analyses pertaining to suppressors of pheS5 were hard to come by. Here we have systematically analyzed a spectrum of Temperature-insensitive derivatives isolated from pheS5 Ts mutant and identified two intragenic suppressors affecting the same base pair coordinate G673 (pheS19 defines G673 → T673; Gly225 → Cys225 and pheS28 defines G673 → C673; Gly225 → Arg225). In fact in the third derivative, the intragenic suppressor originally named pheS43 (G673 → C673transversion) is virtually same as pheS28. In the fourth case, the very pheS5 lesion itself has got changed from A293 → T293 (named pheS40). Cloning of pheS+, pheS5, pheS5-pheS19, pheS5-pheS28 alleles into pBR322 and introduction of these clones into pheS5 mutant revealed that excess of double mutant protein is not at all good for the survival of cells at 42°C. These results clearly indicate a pivotal role for Gly225 in the structural/functional integrity of alpha subunit of E. coli PheRS enzyme and it is proposed that G673 might define a hot spot for intragenic suppressors of pheS5.

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Allele-specific suppression of the temperature sensitivity offitA/fitB mutants ofEscherichia coli by a new mutation (fitC4): Isolation, characterization and its implications in transcription control

et al. 2006

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The temperature sensitive transcription defective mutant of Escherichia coli originally called fitA76 has been shown to harbour two missense mutations namely pheS5 and fit95. In order to obtain a suppressor of fitA76, possibly mapping in rpoD locus, a Ts+ derivative (JV4) was isolated from a fitA76 mutant. It was found that JV4 neither harbours the lesions present in the original fitA76 nor a suppressor that maps in or near rpoD. We show that JV4 harbours a modified form of fitA76 (designated fitA76*) together with its suppressor. The results presented here indicate that the fit95 lesion is intact in the fitA76* mutant and the modification should be at the position of pheS5. Based on the cotransduction of the suppressor mutation and/or its wild type allele with pps, aroD and zdj-3124::Tn10 kan we have mapped its location to 39.01 min on the E. coli chromosome. We tentatively designate the locus defined by this new extragenic suppressor as fitC and the suppressor allele as fitC4. While fitC4 could suppress the Ts phenotype of fitA76* present in JV4, it fails to suppress the Ts phenotype of the original fitA76 mutant (harbouring pheS5 and fit95). Also fitC4 could suppress the Ts phenotype of a strain harbouring only pheS5. Interestingly, the fitC4 Ts phenotype could also be suppressed by fit95. The pattern of decay of pulse labelled RNA in the strains harbouring fitC4 and the fitA76* resembles that of the original fitA76 mutant implying a transcription defect similar to that of fitA76 in both these mutants. The implications of these findings with special reference to transcription control by Fit factors in vivo are discussed.

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Elucidation of the lesions present in the transcription defectivefitA76 mutant ofEscherichia coli: Implication of phenylalanyl tRNA synthetase subunits as transcription factors

Cited by 6 publications

References 23 publications

Glu₅₇₁ of PheT plays a pivotal role in the thermal stability of Escherichia coli PheRS enzyme

Glu₅₇₁ of PheT plays a pivotal role in the thermal stability of Escherichia coli PheRS enzyme

G₆₇₃ could be a novel mutational hot spot for intragenic suppressors of pheS5 lesion in Escherichia coli

Allele-specific suppression of the temperature sensitivity offitA/fitB mutants ofEscherichia coli by a new mutation (fitC4): Isolation, characterization and its implications in transcription control

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Elucidation of the lesions present in the transcription defectivefitA76 mutant ofEscherichia coli: Implication of phenylalanyl tRNA synthetase subunits as transcription factors

Cited by 6 publications

References 23 publications

Glu571 of PheT plays a pivotal role in the thermal stability of Escherichia coli PheRS enzyme

Glu571 of PheT plays a pivotal role in the thermal stability of Escherichia coli PheRS enzyme

G673 could be a novel mutational hot spot for intragenic suppressors of pheS5 lesion in Escherichia coli

Allele-specific suppression of the temperature sensitivity offitA/fitB mutants ofEscherichia coli by a new mutation (fitC4): Isolation, characterization and its implications in transcription control

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Glu₅₇₁ of PheT plays a pivotal role in the thermal stability of Escherichia coli PheRS enzyme

Glu₅₇₁ of PheT plays a pivotal role in the thermal stability of Escherichia coli PheRS enzyme

G₆₇₃ could be a novel mutational hot spot for intragenic suppressors of pheS5 lesion in Escherichia coli