Substrate recognition mechanism of tRNA-targeting ribonuclease, colicin D, and an insight into tRNA cleavage-mediated translation impairment.
RNA Biol
; 18(8): 1193-1205, 2021 08.
Article
en En
| MEDLINE
| ID: mdl-33211605
ABSTRACT
Colicin D is a plasmid-encoded bacteriocin that specifically cleaves tRNAArg of sensitive Escherichia coli cells. E. coli has four isoaccepting tRNAArgs; the cleavage occurs at the 3' end of anticodon-loop, leading to translation impairment in the sensitive cells. tRNAs form a common L-shaped structure and have many conserved nucleotides that limit tRNA identity elements. How colicin D selects tRNAArgs from the tRNA pool of sensitive E. coli cells is therefore intriguing. Here, we reveal the recognition mechanism of colicin D via biochemical analyses as well as structural modelling. Colicin D recognizes tRNAArgICG, the most abundant species of E. coli tRNAArgs, at its anticodon-loop and D-arm, and selects it as the most preferred substrate by distinguishing its anticodon-loop sequence from that of others. It has been assumed that translation impairment is caused by a decrease in intact tRNA molecules due to cleavage. However, we found that intracellular levels of intact tRNAArgICG do not determine the viability of sensitive cells after such cleavage; rather, an accumulation of cleaved ones does. Cleaved tRNAArgICG dominant-negatively impairs translation in vitro. Moreover, we revealed that EF-Tu, which is required for the delivery of tRNAs, does not compete with colicin D for binding tRNAArgICG, which is consistent with our structural model. Finally, elevation of cleaved tRNAArgICG level decreases the viability of sensitive cells. These results suggest that cleaved tRNAArgICG transiently occupies ribosomal A-site in an EF-Tu-dependent manner, leading to translation impairment. The strategy should also be applicable to other tRNA-targeting RNases, as they, too, recognize anticodon-loops.Abbreviations mnm5U 5-methylaminomethyluridine; mcm5s2U 5-methoxycarbonylmethyl-2-thiouridine.
Palabras clave
Texto completo:
1
Bases de datos:
MEDLINE
Asunto principal:
Ribosomas
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Bacteriocinas
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Biosíntesis de Proteínas
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ARN Bacteriano
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ARN de Transferencia de Arginina
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Colicinas
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Escherichia coli
Tipo de estudio:
Prognostic_studies
Idioma:
En
Revista:
RNA Biol
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
2021
Tipo del documento:
Article
País de afiliación:
Japón