Correct codon-anticoden base pairing at the 5'-anticodon position blocks covalent crosslinking between transfer ribonucleic acid and 16S RNA at the ribosomal P site

Ofengand, James; Liou, Richard

doi:10.1021/bi00506a017

Cited by 37 publications

(37 citation statements)

References 25 publications

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“…E. coli N-acetylvalyl (AcVal)-tRNAv"1, E. coli tight couple ribosomes, poly(U2,G), pG-U-U, G-U-A, and Sephacryl S-200 were prepared or obtained as described (15,16). Poly(U5,G) was from P-L Biochemicals.…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Functional conservation near the 3' end of eukaryotic small subunit RNA: photochemical crosslinking of P site-bound acetylvalyl-tRNA to 18S RNA of yeast ribosomes.

Ofengand¹,

Górnicki²,

Chakraburtty³

et al. 1982

Proc. Natl. Acad. Sci. U.S.A.

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Escherichia coli acetylvalyl (AcVal)-tRNAva' became crosslinked to both yeast and spinach chloroplast ribosomes upon irradiation (300 nm) in the presence of poly(U2,G). Yields were 25-30% and 33%, respectively, compared to 45% for E. coli. Crosslinking occurred to the P site, only to the 40S subunit, and 90% of that was to the 18S rRNA. The crosslink could be photolyzed at 254 nm with the same first-order kinetics as for the E. coli ribosome complex previously studied. The AcVal-tRNA that split off could be crosslinked again when irradiated at 300 nm, showing that the crosslink was photoreversible. There was a strong codon specificity for crosslinking. With pG

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Section: Methodsmentioning

confidence: 99%

“…Base-pairing of the crosslinking residue, (c)mo5U-34, in the tRNA with either A or G in the codon also blocked crosslinking (16). The structure of the crosslink was shown to be a pyrimidine-pyrimidine cyclobutane dimer between the (c)mo5U-34 residue of the tRNA and a pyrimidine of the 16S RNA (17).…”

mentioning

confidence: 97%

Functional conservation near the 3' end of eukaryotic small subunit RNA: photochemical crosslinking of P site-bound acetylvalyl-tRNA to 18S RNA of yeast ribosomes.

Ofengand¹,

Górnicki²,

Chakraburtty³

et al. 1982

Proc. Natl. Acad. Sci. U.S.A.

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“…This crosslinking was done with an empty A site, so C-1400 would be available. When the proper codon was supplied for the tRNA (Ofengand and Liou, 1981) crosslinking was abolished. This indicates that the interaction between the anticodon and C-1400 is probably not functionally important.…”

Section: Proofreading and Trna Bindingmentioning

confidence: 99%

Structure-function relations in E. coli 16S RNA

Thompson¹,

Hearst²

1983

Cell

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“…In view of this correlation, we suggested that the 5' anticodon base is the photoreactive moiety in the tRNA (2,3). Moreover, the efficiency of crosslinking was found to vary widely, depending on the residue in the wobble position (2,3) and its ability to base-pair with the corresponding nucleotide in the codon (7). Despite such strong circumstantial evidence, it has not yet been demonstrated directly that the modified uridine of the anticodon is the crosslinked residue.…”

mentioning

confidence: 99%

Covalent crosslinking of tRNA1Val to 16S RNA at the ribosomal P site: identification of crosslinked residues.

Prince¹,

Taylor²,

Thurlow³

et al. 1982

Proc. Natl. Acad. Sci. U.S.A.

199

107

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N-Acetylvalyl-tRNAYl (AcVal-tRNA 1) was bound to the P site of uniformly 32P-labeled 70S ribosomes from Escherichia coli and crosslinked to 16S RNA in the 30S ribosomal subunit by irradiation with light of300-400 nm. To identify the crosslinked nucleotide in 16S RNA, AcVal-tRNAY. l-16S [32P]RNA was digested completely with RNase TI and the band containing the covalently attached oligonucleotides from tRNA and rRNA was isolated by polyacrylamide gel electrophoresis. The crosslinked oligonucleotide, and the 32P-labeled rRNA moiety released from it by photoreversal of the crosslink at 254 nm, were then analyzed by secondary hydrolysis with pancreatic RNase A and RNase U2. The oligonucleotide derived from 16S RNA was found to be the evolutionarily conserved sequence, U-A-C-A-C-A-C-C-G1401, and the nucleotide crosslinked to tRNAI , C14W. The identity of the covalently attached residue in the tRNA was established by using AcVal-tRNAlv-16S RNA prepared from unlabeled ribosomes.This complex was digested to completion with RNase T1 and the resulting RNA fragments were labeled at the 3' end with [5'-32P]pCp. The crosslinked Ti oligonucleotide isolated from the mixture yielded one major end-labeled component upon photoreversal. Chemical sequence analysis demonstrated that this product was derived from the anticodon-containing pentadecanucleo-

show abstract

Correct codon-anticoden base pairing at the 5'-anticodon position blocks covalent crosslinking between transfer ribonucleic acid and 16S RNA at the ribosomal P site

Cited by 37 publications

References 25 publications

Functional conservation near the 3' end of eukaryotic small subunit RNA: photochemical crosslinking of P site-bound acetylvalyl-tRNA to 18S RNA of yeast ribosomes.

Functional conservation near the 3' end of eukaryotic small subunit RNA: photochemical crosslinking of P site-bound acetylvalyl-tRNA to 18S RNA of yeast ribosomes.

Structure-function relations in E. coli 16S RNA

Covalent crosslinking of tRNA1Val to 16S RNA at the ribosomal P site: identification of crosslinked residues.

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