The primary structure of protein L27 from the peptidyl-tRNA binding site of Escherichia coli ribosomes.
TL;DR: The ribosomal protein L27 is a constituent of the larger (50S) subunit of the E. coil ribosome and can be inferred to be very close to the peptidyl-tRNA binding site (P-site).
About: This article is published in FEBS Letters.The article was published on 1975-11-01 and is currently open access. It has received 34 citations till now. The article focuses on the topics: TRNA binding & Ribosome.
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2,751 citations
TL;DR: These structures demonstrate that the PTC is very similar between the 50S subunit and the intact ribosome, and reveal interactions between the ribosomal proteins L16 and L27 and the tRNA substrates, helping to elucidate the role of these proteins in peptidyl transfer.
Abstract: Protein synthesis is catalyzed in the peptidyl transferase center of the ribosome. The structure of the 70S ribosome containing tRNAs now gives insight into the active site of a complete ribosome and reveals a direct interaction between the tRNA substrate and ribosomal proteins. Protein synthesis is catalyzed in the peptidyl transferase center (PTC), located in the large (50S) subunit of the ribosome. No high-resolution structure of the intact ribosome has contained a complete active site including both A- and P-site tRNAs. In addition, although past structures of the 50S subunit have found no ordered proteins at the PTC, biochemical evidence suggests that specific proteins are capable of interacting with the 3′ ends of tRNA ligands. Here we present structures, at 3.6-A and 3.5-A resolution respectively, of the 70S ribosome in complex with A- and P-site tRNAs that mimic pre- and post-peptidyl-transfer states. These structures demonstrate that the PTC is very similar between the 50S subunit and the intact ribosome. They also reveal interactions between the ribosomal proteins L16 and L27 and the tRNA substrates, helping to elucidate the role of these proteins in peptidyl transfer.
352 citations
TL;DR: The cross‐linked amino acids in ribosomal proteins S17 and L6 prove the proposed RNA interacting site derived from three‐dimensional models, and a comparison of the structural data with mutations in ribOSomal proteins that lead to antibiotic resistance, and with those from protein‐antibiotic cross‐linking experiments, reveals functional implications for ribosome proteins that interact with rRNA.
Abstract: We have investigated protein-rRNA cross-links formed in 30S and 50S ribosomal subunits of Escherichia coli and Bacillus stearothermophilus at the molecular level using UV and 2-iminothiolane as cross-linking agents. We identified amino acids cross-linked to rRNA for 13 ribosomal proteins from these organisms, namely derived from S3, S4, S7, S14, S17, L2, L4, L6, L14, L27, L28, L29 and L36. Several other peptide stretches cross-linked to rRNA have been sequenced in which no direct cross-linked amino acid could be detected. The cross-linked amino acids are positioned within loop domains carrying RNA binding features such as conserved basic and aromatic residues. One of the cross-linked peptides in ribosomal protein S3 shows a common primary sequence motif--the KH motif--directly involved in interaction with rRNA, and the cross-linked amino acid in ribosomal protein L36 lies within the zinc finger-like motif of this protein. The cross-linked amino acids in ribosomal proteins S17 and L6 prove the proposed RNA interacting site derived from three-dimensional models. A comparison of our structural data with mutations in ribosomal proteins that lead to antibiotic resistance, and with those from protein-antibiotic cross-linking experiments, reveals functional implications for ribosomal proteins that interact with rRNA.
121 citations
TL;DR: The results demonstrate that deamidation can alter points of proteolytic cleavage of human growth hormone and may be a way of directing specific cleavages.
104 citations
TL;DR: The results indicate that MRP13 expression is regulated predominantly at the transcriptional level in response to catabolite repression and the cellular capacity for respiration and, in addition, that protein levels appear to be modulated posttranscriptionally by degradation of free copies of the MRp13 protein.
Abstract: MRP13 is defined by biochemical criteria as a 35-kilodalton small subunit protein of the yeast mitochondrial ribosome. The MRP13 gene was identified by immunological screening of a yeast genomic library in lambda gt11 and a functional copy of the gene has been cloned on a 2.2-kilobase BglII fragment. Sequencing of this fragment showed that the MRP13 coding region specifies a 324-amino-acid basic protein with a calculated Mr of 37,366. Computer searches failed to reveal any significant sequence similarity to previously identified ribosomal proteins or to the sequences in the current National Biomedical Research Foundation data base. Cells carrying disrupted copies of MRP13 lacked the MRP13 protein but were not impaired in either mitochondrial protein synthesis or assembly of 37S ribosomal subunits, indicating that, like L29 and L30 in Escherichia coli (M. Lotti, E. R. Dabbs, R. Hasenbank, M. Stoffler-Meilicke, and G. Stoffler, Mol. Gen. Genet. 192:295-300, 1983), MRP13 is not essential for ribosome synthesis or function. Analysis of the sequence in the MRP13 5'-flanking region revealed the closely linked gene for the cytoplasmic ribosomal protein rp39A. The rp39A coding region began at nucleotide -846 and ended at -325 with respect to the MRP13 translational start. The steady-state levels of the MRP13 mRNA were determined in response to carbon catabolite repression, variation in the mitochondrial genetic background, and increased gene dosage of MRP13. In [rho+] cells, transcript levels were repressed severalfold by growth in glucose compared with growth in either galactose or nonfermentable carbon sources. In respiratory-deficient strains ([rho0], [mit-]), however, transcription appeared to be largely derepressed even in the presence of high concentrations of glucose. Despite high levels of the MRP13 transcripts in [rho0] cells, the MRP13 protein did not accumulate, suggesting that the protein is relatively unstable in the absence of ribosome assembly. Cells carrying the MRP13 gene on a multiple-copy plasmid overproduced the mRNA in rough proportion to the gene dosage and the protein in a significant but lesser amount. The results indicate that MRP13 expression is regulated predominantly at the transcriptional level in response to catabolite repression and the cellular capacity for respiration and, in addition, that protein levels appear to be modulated posttranscriptionally by degradation of free copies of the MRP13 protein.
89 citations
References
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TL;DR: The helix, s Applequist, 1963) in which the Zimm-Bragg parameters u and s are defined respectively as the cooperativity factor for helix initiation, and the equi- librium constant for converting a coil residue to a helical helix.
Abstract: The helix, s Applequist, 1963) in which the Zimm-Bragg parameters u and s are defined respectively as the cooperativity factor for helix initiation, and the equi- librium constant for converting a coil residue to a helical ~~~~
2,112 citations
690 citations
TL;DR: The staphylococcal protease hydrolyzes all of the seventeen different glutamoyl bonds studied, although those involving hydrophobic aminoacid residues with bulky side chains are cleaved at a lower rate.
Abstract: An extracellular protease of Staphylococcus aureus, strain V8, previously shown to cleave specifically the peptide bonds on the carboxyl-terminal side of either aspartate or glutamate residues in phosphate buffer (pH 7.8) hydrolyzes only glutamoyl bonds in either ammonium bicarbonate (pH 7.8) or ammonium acetate (pH 4.0). Of all aspartoyl bonds tested, only the Asp-Gly linkage is cleaved at a detectable rate. The staphylococcal protease hydrolyzes all of the seventeen different glutamoyl bonds studied, although those involving hydrophobic aminoacid residues with bulky side chains are cleaved at a lower rate.
640 citations
TL;DR: Investigation of whether the use of maleylation to introduce glyoxyloyl groups (Dixon, 1968) could be extended to the introduction of pyruvoyl groups reports the effects of introducing methyl groups into the molecule of maleic anhydride.
Abstract: Butler, Harris, Hartley & Leberman (1967) showed that maleic anhydride could be used for the reversible blocking of amino groups. The maleyl group could be removed because the protonated form of the free carboxyl group catalysed the hydrolysis of the amide bond, presumably by intramolecular general acid catalysis (cf. Bender, Chow & Chloupek, 1958). The present paper reports the effects of introducing methyl groups into the molecule of maleic anhydride. 2,3-Dimethylmaleic anhydride. We had wished to investigate whether the use of maleylation to introduce glyoxyloyl groups (Dixon, 1968) could be extended to the introduction of pyruvoyl groups. We therefore treated arginine with 2,3-dimethylmaleic anhydride (Fluka A.-G., Buchs, Switzerland) as follows. To a solution of arginine hydrochloride (5M) in water was added, with stirring, dimethylmaleic anhydride to a final concentration of 5-5m, and the pH was maintained at 8 by the addition of N-NaOH. Uptake ofbase ceased after about an hour and suitable samples were then applied to paper for high-voltage electrophoresis in the buffer systems used previously (Perham, 1967). Material was detected on the paper by means of the ninhydrincadmium reagent (Heilmann, Barrollier & Watzke, 1957) and also by the Sakaguchi reaction (Jepson & Smith, 1953). It was immediately apparent that arginine alone and no product could be detected after electrophoresis at pH 3 5, but that at pH 6 5
394 citations