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- Nucleotide Sequence and Secondary Structure of 5S rRNA from Sphingobium chungbukense DJ77
- Hae-Ryong Kwon , Young-Chang Kim
- J. Microbiol. 2007;45(1):79-82.
- DOI: https://doi.org/2486 [pii]
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Abstract
- The 5S rRNA gene from Sphingobium chungbukense DJ77 was identified. The secondary structure of the 199-base-long RNA was proposed. The two-base-long D loop was the shortest among all of the known 5S rRNAs. The U19-U64 non-canonical pair in the helix II region was uniquely found in strain DJ77 among all of the sphingomonads.
- The Influence of the Nucleotide Sequences of Random Shine-Dalgarno and Spacer Region on Bovine Growth Hormone Gene Expression
- Soon-Young Paik , Kyung Soo Ra , Hoon Sik Cho , Kwang Bon Koo , Hyung Suk Baik , Myung Chul Lee , Jong Won Yun , Jang Won Choi
- J. Microbiol. 2006;44(1):64-71.
- DOI: https://doi.org/2335 [pii]
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Abstract
- To investigate the effects of the nucleotide sequences in Shine-Dalgarno (SD) and the spacer region (SD-ATG) on bovine growth hormone (bGH) gene expression, the expression vectors under the control of the T7 promoter (pT7-7 vector) were constructed using bGH derivatives (bGH1 & bGH14) which have different 5''-coding regions and were induced in E. coli BL21(DE3). Oligonucleotides containing random SD sequences and a spacer region were chemically synthesized and the distance between the SD region and the initiation codon were fixed to nine bases in length. The oligonucleotides were annealed and fused to the bGH1 and bGH14 cDNA, respectively. When the bGH gene was induced with IPTG in E. coli BL21(DE3), some clones containing only bGH14 cDNA produced considerable levels of bGH in the range of 6.9% to 8.5% of total cell proteins by SDS-PAGE and Western blot. Otherwise, the bGH was not detected in any clones with bGH1 cDNA. Accordingly, the nucleotide sequences of SD and the spacer region affect on bGH expression indicates that the sequences sufficiently destabilize the mRNA secondary structure of the bGH14 gene. When the free energy was calculated from the transcription initiation site to the +51 nucleotide of bGH cDNA using a program of nucleic acid folding and hybridization prediction, the constructs with values below ‒26.3 kcal/mole (toward minus direction) were not expressed. The constructs with the original sequence of bGH cDNA also did not show any expression, regardless of the free energy values. Thus, the disruption of the mRNA secondary structure may be a major factor regulating bGH expression in the translation initiation process. Accordingly, the first stem-loop among two secondary structures present in the 5''-end region of the bGH gene should be disrupted for the effective expression of bGH.
- Nucleotide Sequence Analysis of the 5S Ribosomal RNA Gene of the Mushroom Tricholoma matsutake
- Hwang, Seon Kap , Kim, Jong Guk
- J. Microbiol. 1995;33(2):136-141.
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Abstract
- From a cluster of structural rRNA genes which has previsouly been cloned (Hwang and Kim, in submission; J. Microbiol. Biotechnol.), a 1.0-kb Eco RI fragment of DNA which shows significant homology to the 25S and rRNA s of Tricholoma matsutake was used for sequence analysis. Nucleotide sequence was bidirectionally determined using deletion series of the DNA fragment. Comparing the resultant 1016-base sequence with sequences in the database, both the 3'end of 25S-rRNA gene and 5S rRNA gene were searched. The 5S rRNA gene is 118-bp in length and is located 158-bp downstream of 3'end of the 25S rRNA gene. IGSI and IGS2 (partial) sequences are also contained in the fragment. Multiple alignment of the 5S rRNA sequences was carried out with 5S rRNA sequences from some members of the subdivision Basidiomycotina obtained from the database. Polygenetic analysis with distance matrix established by Kimura's 2-parameter method and phylogenetic tree by UPGMA method proposed that T. matsutake is closely related to efibulobasidium allbescens. Secondary structure of 5S rRNA was also hypothesized to show similar topology with its generally accepted eukaryotic counterpart.
- Secondary Structure Analysis of Amino Terminal Domain in Phage Lambda Integrase
- Yu, Jeong A , Nam, Chan Eun , Cho, Eun Hee
- J. Microbiol. 1998;36(4):266-272.
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Abstract
- The amino-terminal domain of bacteriophage λ integrase recognizes specific DNA sequences called arm-type sites. To study the structural and functional relationships of the integras armtype DNA binding domains were confirmed by gel mobility-shift assay. The polypeptides were subjected to circular dichroism spectroscopy to estimate the amount of secondary structures they contain Based upon analyses of circular dichroism spectra and comparison with predicted secondary structural compositions, it was estimated that the amino terminal domain of integrase in an aqueous solution was composed of a little α-helical region. The helical content increased with an increasing amount of ethanol, an α-helix inducer. This indicates that its conformation can be changed to a form with higher content of α-helical structure under a certain condition.
- Continuous Synthesis of Escherichia coli GroEL at a High Temperature
- Young Hak Kwak , Kyong Sun Lee , Ji Yeon Kim , Dong Seok Lee , Han Bok Kim
- J. Microbiol. 2000;38(3):145-149.
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Abstract
- GroEL is a typical molecular chaperone. GroEL synthesis patterns at various culture temperatures in Escherichia coli were investigated in this study. No significant differences in the amount of GroEL produced from the chromosome were found at 30 and 37 C. However, GroEL production increased 3.4-fold at 42 C. GroEL synthesis was not transient but continuous at 42 C, although most heat shock gene expression is known to be transient. To understand the role of the groEL structural gene, a groE promoter-lacZ fusion was constructed. Interestingly, while transcriptional fusion is not thermally inducible, it is inducible by ethanol, suggesting that the secondary structure of the groEL transcript is involved in thermal regulation of the groEL gene. Secondary structures of groE mRNA at 37 and 42 C were compared using the computer program RNAdraw. Distinct structures at the two temperatures were found, and these structures may be related to a high level of GroEL expression at 42 C.
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