Role of the aminoacyl end of transfer RNA in the allosteric control of guanosine pentaphosphate synthesis by the stringent factor-ribosome complex of Escherichia coli

Chinali, Gianni; Liou, Richard; Ofengand, James

doi:10.1021/bi00607a010

Cited by 9 publications

(10 citation statements)

References 26 publications

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“…Such a situation can only be reached by tRNAs without amino acid, and hence not fully engaged on the A-site. Previous data demonstrated the critical importance of a free CCA end in the activation of the factor [10][11][12]. Indeed, binding experiments using the nonhydrolizable Phe-NH-tRNA Phe (supplementary Fig S4 online) show that in vitro the presence of deacylated tRNA promotes the maximum binding of RelA to the ribosome.…”

Section: Discussionmentioning

confidence: 87%

The ribosome triggers the stringent response by RelA via a highly distorted tRNA

et al. 2013

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The bacterial stringent response links nutrient starvation with the transcriptional control of genes. This process is initiated by the stringent factor RelA, which senses the presence of deacylated tRNA in the ribosome as a symptom of amino-acid starvation to synthesize the alarmone (p)ppGpp. Here we report a cryo-EM study of RelA bound to ribosomes bearing cognate, deacylated tRNA in the A-site. The data show that RelA on the ribosome stabilizes an unusual distorted form of the tRNA, with the acceptor arm making contact with RelA and far from its normal location in the peptidyl transferase centre.

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Section: Discussionmentioning

confidence: 87%

The ribosome triggers the stringent response by RelA via a highly distorted tRNA

et al. 2013

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“…To explore this speculation it will be necessary to examine to see if 3' fluorescent derivatives can mimic deacylated tRNA in triggering the stringent response. Offengand and coworkers (32,33) have localized the effect of deacylated tRNA to the terminal adenosine at the 3' end but the effects of particular 3' derivatives remain unknown.…”

Section: Discussionmentioning

confidence: 99%

Ribosome binding by tRNAs with fluorescent labeled 3′ termini

Wells

Cantor

1980

Nucl Acids Res

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Yeast and E. coli tRNAPhe samples were oxidized and labeled at the 3' end with dansyl hydrazine or fluorescein thiosemicarbazide. These tRNAs can bind to poly(U)-programmed E. coli 70S tight couple ribosomes in 25 mM magnesium at 8 degrees C. Two binding sites with binding constants of about 1 X 10(9) M-1 (P) and 3 X 10(7) M-1 (A) were determined for the yeast tRNAPhe derivatives. With E. coli tRNAPhe the A site affinity is similar to yeast tRNAPhe but the P site affinity is 5-fold weaker. Singlet-singlet energy transfer showd that the distance from the 3' end of tRNAPhe in the P site to a fluorescein derivative of erythromycin is 23 A. This supports in vitro studies suggesting that erythromycin binds near the peptide moiety of peptidyl tRNA. A distance of 34 A between the 3' ends of 2 tRNAs bound simulatneously on the ribosome was also measured. This long distance may mean that the deacylated fluorescent tRNA binds to the A site in an orientation like that in the stringent response rather than in protein synthesis.

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“…High salt washed "loose couple" ribosomes, stringent factor, and phenylalanyl-tRNA synthetase were prepared as described (3,4) as were APAA-Br and [ H]NAG-NOS (7). Methods.…”

Section: Methodsmentioning

confidence: 99%

“…Under appropriate conditions, the rate of synthesis is directly proportional to the concentration of tRNA added (2)(3)(4)(5). Chemically modified tRNA' species vary in their activity in this system both with respect to their relative affinities for the ribosome-stringent factor complex and in the maximum rate of synthesis that they can induce (4). Consequently, a study of the activity of modified tRNAs can help to delineate those features of the tRNA molecule which are recognized by stringent factor.…”

Section: Introductionmentioning

confidence: 99%

“…Consequently, a study of the activity of modified tRNAs can help to delineate those features of the tRNA molecule which are recognized by stringent factor. Previous studies have shown that none of the uridine or uridine-derived nucleotides (such as pseudouridine) are essential, since they can all be replaced by 5fluorouridine (3), but that the exact structure of the 3'-end is very important (4,5). Except for a preliminary report that substitution of uridine for ribothymidine had no effect, and that replacement of N,N-dimethyl guano-sine by guanosine did inactivate (6), no other tRNA modifications have been studied.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Ability of modified forms of phenylalanine tRNA to stimulate guanosine pentaphosphate synthesis by the stringent factor-ribosome complex of E. coli

Ofengand

Liou

1978

Nucl Acids Res

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PRNhe 4 tRNAPhe of E. coli, modified at its 4-thiouridine ( Srd) and 3-(3-amino-3-carboxypropyl)uridine (nbt3UXd) residues, was tested for its ability to induce (p)ppGpp synthesis. The Srd residue was derivatized with the p-azidophenacyl group, cross-linked to Cyd13, and the borohydride reduction'product of the cross-link was prepared. The nbt3Urd residue was derivatized with the N-(4-azido-2-nitrophenyl)glycyl group. None of these derivatives had more than a minor effect on the affinity of the tRNA for the stringent factorribosome complex, and no effect at all on the maximum velocity of (p)ppGpp synthesis, either at 2 or 82 mM NH4C1. These two regions of the tRNA which are on opposite faces of the tRNA molecule do not appear to be structurally important for recognition by the stringent factor-ribosome complex. They may provide useful sites, therefore, for the introduction of photoaffinity or fluorescent probes with which to study tRNA-stringent factor recognition.

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Role of the aminoacyl end of transfer RNA in the allosteric control of guanosine pentaphosphate synthesis by the stringent factor-ribosome complex of Escherichia coli

Cited by 9 publications

References 26 publications

The ribosome triggers the stringent response by RelA via a highly distorted tRNA

The ribosome triggers the stringent response by RelA via a highly distorted tRNA

Ribosome binding by tRNAs with fluorescent labeled 3′ termini

Ability of modified forms of phenylalanine tRNA to stimulate guanosine pentaphosphate synthesis by the stringent factor-ribosome complex of E. coli

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