• Bulletin of the Korean Chemical Society
• /
• v.20 no.11
• /
• pp.1335-1339
• /
• 1999

The function of 5S rRNA, a constituent of a large subunit of ribosome, is not clearly known yet. To identify RNA motifs interacting with 5S rRNA, and thereby to get an insight into the function of 5S rRNA in the ribosome, a SELEX (Systematic Evolution of Ligands by Exponential Enrichment) experiment was performed. RNA molecules binding to Escherichia coli 5S rRNA were selected from a 48-mer random sequence library through 12 rounds of selection, cloned, and sequenced. Two groups of the selected RNA molecules had the consensus sequences GCGG and GUGAAA, respectively, which are present in the segment, G688 through A696, of E. coli 16S rRNA. The gel mobility shift assay showed that 5S rRNA interacted with the 16S rRNA fragment containing the GCGG and GUGAAA sequences. The enzymatic protection experiment shows that the A29CCUGA34 and G51AAGUG56 sequences of 5S rRNA and the C680AGG683 and G688CGG691 sequences of the 16S rRNA fragment are involved in the interaction between the two RNA molecules. On the basis of this observation, we suggest that 5S rRNA and 16S rRNA play a role for the association of two ribosomal subunits.

• BMB Reports
• /
• v.52 no.8
• /
• pp.502-507
• /
• 2019

Translation is a costly, but inevitable, cell maintenance process. To reduce unnecessary ATP consumption in cells, a fine-tuning mechanism is needed for both ribosome biogenesis and translation. Previous studies have suggested that the ribosome functions as a hub for many cellular signals such as ribotoxic stress response, mammalian target of rapamycin (mTOR), and ribosomal S6 kinase (RSK) signaling. Therefore, we investigated the relationship between ribosomes and mitogen-activated protein kinase (MAPK) activation under ribotoxic stress conditions and found that the activation of c-Jun N-terminal kinases (JNKs) was suppressed by ribosomal protein knockdown but that of p38 was not. In addition, we found that JNK activation is driven by the association of inactive JNK in the 80S monosomes rather than the polysomes. Overall, these data suggest that the activation of JNKs by ribotoxic stress is attributable to 80S monosomes. These 80S monosomes are active ribosomes that are ready to initiate protein translation, rather than polysomes that are already acting ribosomes involved in translation elongation.

• Microbiology and Biotechnology Letters
• /
• v.39 no.1
• /
• pp.93-96
• /
• 2011

It has been shown that a nucleotide substitution at position 770 in Escherichia coli 16S rRNA, which is implicated in forming the evolutionary conserved B2c intersubunit bridge, has a detrimental effect on ribosome function. In order to isolate second-site revertants that complement ribosomes containing C770G, we performed a random mutagenesis of the 16S rRNA gene and selected clones that could produce more CAT protein translated by specialized ribosome. One of the clones contained two nucleotide substitutions at positions 569 and 904 (C569G and U904C) and these mutations partially complemented the loss of protein-synthesis ability caused by C770G. Further studies using the isolated revertant will provide information about which part of 16S rRNA is interacting with C770 and the consequence of the structure formed by these interactions in the process of protein synthesis.

• Journal of Life Science
• /
• v.32 no.3
• /
• pp.249-255
• /
• 2022

Metabolism is essential for survival and reproduction, and there is a metabolic pathways entry in the clusters of orthologous groups of proteins (COGs) database, updated in 2020. In this study, the metabolic pathways of 1309 prokaryotes were analyzed using COGs. There were 822 COGs associated with 63 metabolic pathways, and the mean for each taxon was between 200.50 (mollicutes) and 527.07 (cyanobacteria) COGs. The metabolic pathway composition ratio (MPCR) was defined as the number of COGs present in one genome in relation to the total number of COGs constituting each metabolic pathway, and the number of pathways with 100% MPCR ranged from 0 to 26 in each prokaryote. Among 1309 species, the 100% MPCR pathways included murein biosynthesis associated with cell wall synthesis (922 species); glycine cleavage (918); and ribosomal 30S subunit synthesis (903). The metabolic pathways with 0% MPCR were those involving photosystem I (1263 species); archaea/vacuolar-type ATP synthase (1028); and Na⁺-translocation NADH dehydrogenase (976). Depending on the prokaryote, three to 49 metabolic pathways could not be performed at all. The sequence of most highly conserved metabolic pathways was ribosome 30S subunit synthesis (96.1% of 1309 species); murein biosynthesis (86.8%); arginine biosynthesis (80.4%); serine biosynthesis (80.3%); and aminoacyl-tRNA synthesis (82.2%). Protein and cell wall synthesis have been shown to be important metabolic pathways in prokaryotes, and the results of this study of COGs related to such pathways can be utilized in, for example, the development of antibiotics and artificial cells.

• Journal of Life Science
• /
• v.32 no.6
• /
• pp.463-467
• /
• 2022

As a result of applying the COG (Cluster of Orthologous Groups of Protein) algorithm to 1,309 species to confirm the conserved genes of prokaryotes, ribosomal protein S11 (COG0100) was identified. The numbers of conservative genes were 2, 5, 5, and 6 in 1,308, 1,307, 1,306, and 1,305 species, respectively. Twenty-nine genes were conserved in over 1,302 species, and they encoded 23 ribosomal proteins, 3 tRNA synthetases, 2 translation factors, and 1 RNA polymerase subunit. Most of them were related to protein production, suggesting the importance of protein expression in prokaryotes. The highest conservative COG was COG0048 (ribosomal protein S12) among the 29 COGs. The 29 conserved genes usually have one protein for each prokaryote. COG0090 (ribosomal protein L2) had not only the lowest conservation value but also the largest standard deviation of phylogenetic distance value. As COG0090 is not only a member of the ribosome, but also a regulator of replication and transcription, it could be inferred that prokaryotes have large variations in COG0090 to survive in various environments. This study could provide data necessary for basic science, tumor control, and development of antibacterial agents.

• Journal of Microbiology and Biotechnology
• /
• v.24 no.1
• /
• pp.8-12
• /
• 2014

Biofilm formation and antibiotic resistance are important determinants for bacterial pathogenicity. Ribonucleases control RNA degradation and there is increasing evidence that they have an important role in virulence mechanisms. In this report, we show that ribonucleases affect susceptibility against ribosome-targeting antibiotics and biofilm formation in Salmonella.

• Proceedings of the Korea Environmental Mutagen Society Conference
• /
• 2001.10b
• /
• pp.189-189
• /
• 2001

• Molecules and Cells
• /
• v.20 no.1
• /
• pp.1-16
• /
• 2005

The peptidyl transferase center (PTC) is located in a protein free environment, thus confirming that the ribosome is a ribozyme. This arched void has dimensions suitable for accommodating the 3'ends of the A-and the P-site tRNAs, and is situated within a universal sizable symmetry-related region that connects all ribosomal functional centers involved in amino-acid polymerization. The linkage between the elaborate PTC architecture and the A-site tRNA position revealed that the A-to P-site passage of the tRNA 3'end is performed by a rotatory motion, which leads to stereochemistry suitable for peptide bond formation and for substrate mediated catalysis, thus suggesting that the PTC evolved by genefusion. Adjacent to the PTC is the entrance of the protein exit tunnel, shown to play active roles in sequence-specific gating of nascent chains and in responding to cellular signals. This tunnel also provides a site that may be exploited for local co-translational folding and seems to assist in nascent chain trafficking into the hydrophobic space formed by the first bacterial chaperone, the trigger factor. Many antibiotics target ribosomes. Although the ribosome is highly conserved, subtle sequence and/or conformational variations enable drug selectivity, thus facilitating clinical usage. Comparisons of high-resolution structures of complexes of antibiotics bound to ribosomes from eubacteria resembling pathogens, to an archaeon that shares properties with eukaryotes and to its mutant that allows antibiotics binding, demonstrated the unambiguous difference between mere binding and therapeutical effectiveness. The observed variability in antibiotics inhibitory modes, accompanied by the elucidation of the structural basis to antibiotics mechanism justifies expectations for structural based improved properties of existing compounds as well as for the development of novel drugs.

• Journal of Microbiology and Biotechnology
• /
• v.25 no.5
• /
• pp.620-628
• /
• 2015

A fungal strain, R9, was isolated from the South Atlantic sediment sample and identified as Aspergillus fumigatus. An antifungal protein, AfAFP_R9, was purified from the culture supernatant of Aspergillus fumigatus R9. AfAFP_R9 was identified to be restrictocin, which is a member of the ribosome-inactivating proteins (RIPs), by MALDI-TOF-TOF-MS. AfAFP_R9 displayed antifungal activity against plant pathogenic Fusarium oxysporum, Alternaria longipes, Colletotrichum gloeosporioides, Paecilomyces variotii, and Trichoderma viride at minimum inhibitory concentrations of 0.6, 0.6, 1.2, 1.2, and 2.4 μg/disc, respectively. Moreover, AfAFP_R9 exhibited a certain extent of thermostability, and metal ion and denaturant tolerance. The iodoacetamide assay showed that the disulfide bridge in AfAFP_R9 was indispensable for its antifungal action. The cDNA encoding for AfAFP_R9 was cloned from A. fumigatus R9 by RT-PCR and heterologously expressed in E. coli. The recombinant AfAFP_R9 protein exhibited obvious antifungal activity against C. gloeosporioides, T. viride, and A. longipes. These results reveal the antifungal properties of a RIP member (AfAFP_R9) from marine-derived Aspergillus fumigatus and indicated its potential application in controlling plant pathogenic fungi.

• Journal of Life Science
• /
• v.15 no.5 s.72
• /
• pp.734-738
• /
• 2005

A gene coding for xylanase from alkali-tolerant Bacillus alcalophilus AX2000 was cloned into Escherichia coli $DH5\alpha$ using pUC19. Among 2,000 transformants, one transformant showed clear zone on the detection agar plate containing oat-spells xylan. Its recombinant plasmid, named pXTY99, was found to carry 7.0 kb insert DNA fragment. When the nucleotide sequence of the cloned xylanase gene (xynT) was determined, xynT gene was found to consist of 1,020 base-pair open reading frame coding for a poly-peptide of 340 amino acids with a deduced molecular weight of 40 kDa. The coding sequence was preceded by a putative ribosome binding site, and the transcription initiation signals. The deduced amino acid sequence of xylanase is similar to those of the xylanases from Bacillus sp. Nl37 and B. stearothermophilus 21 with $61\%$ and $59\%$ identical residues, respectively.