• Korean Journal of Microbiology
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• v.41 no.2
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• pp.125-128
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• 2005

A 16S ribosomal RNA gene from S. chungbukensis DJ77 has been sequenced. This sequence had a length of 1,502 bp and was extended for 29 bp at 5' and for 37 bp at 3' from the partial sequence (1,435 bp) registered in 2000 year. Besides, 1 bp was newly added near to the 3' end. We made the secondary structure of the 16S rRNA based on E. coli model and found four specific regions. We found constant and variable regions in genus Sphingomonas as the result of multiple alignment of 16S rRNA gene sequences from Sphingomonas spp. and S. chungbukensis DJ77. We found a stem loop structure in S. chungbukensis DJ77, which was only discovered in C. jejuni to date. It showed the structural agreement despite the difference of the sequences from the both organisms. Finally, S. chungbukensis DJ77 belonged to cluster II (Sphingobium) group, after the classification using phylogenetic analysis and nucleotide signature analysis.

• Korean Journal of Microbiology
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• v.37 no.2
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• pp.120-123
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• 2001

Sphingomonas chungbukensis DJ77 is able to use phenanthrene and biphenyl as the sole carbon and energy source. Mitomycin C curing experiment suggested that polyaromatic hydrocarbon (PAH) utilization in strain DJ77 was plasmid-encoded. The plasmid cured strains were failed to grow on the minimal medium sprayed with biphenyl or phenanthrene. This was evident from southern hybridizations using a previously cloned DNA segment as a probe. There were positive signals in the palsmid DNA of the wild-type strain DJ77 and the absence of hybridizations with chromosomal DNA from the plasmid DNA of the wild-type strain DJ77 and the absence of hybridizations with chromosomal DNA from the palsmid-cured mutant strains.

• Korean Journal of Microbiology
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• v.41 no.2
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• pp.93-98
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• 2005

The sphingosine kinase gene, which is 969-nucleotide long, was identified during the whole genome sequencing of Sphingomonas chungbukensis DJ77. The amino acid sequence showed the identity of $55\%$ with that of Zymomonas mobilis subsp. mobilis ZM4. C2, C3, and C5 domains of eukaryotic sphingosine kinase were found in sphingosine kinase from Sphingomonas chungbukensis DI77. One of these three conserved sites, GGDG, was predicted as a ATP-binding site, and the functions of the others were unknown currently. The phylogenetic tree constructed by ClustalX indicated that the sphingosine kinase of S. chungbukensis DJ77 was near the phylogenetic group COG1597, and did not belong to the group of diacylglycerol kinase of the same strain. The recombinant sphingosine kinase was expressed in Escherichia coli, but it was made in form of inclusion body.

• Korean Journal of Microbiology
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• v.41 no.1
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• pp.8-12
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• 2005

Some genes, which are involved in the biosynthesis of polysaccharides, could be found by the genome project of Sphingomonas chungbukensis DJ77. In this study, we identified the complete nucleotide sequence of a gene, encoding the glucosyl-isoprenyl phosphate-transferase, which catalyzes the first step in the biochemical pathway for the synthesis of the sphingan type polysaccharide. This gene, named spsB, is initiated by the ATG codon and terminated by the TGA, and its open reading frame consists of 1392 bp, encoding 463 amino acids. The predicted amino acid sequence of this enzyme indicates $50\%$ similarity to SpsB of Sphingomonas spp S88, also produces sphingan, and $48\%$ to GelB of Sphingomonas paucimobilis ATCC 31461.

• Korean Journal of Microbiology
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• v.39 no.3
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• pp.123-127
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• 2003

Sphingomonas chungbukensis DJ77 is able to metabolize phenanthrene as the sole carbon and energy source. The plasmid pUPX5 includes phnS gene encoding 2-hydroxychromene-2-carboxylate (HCCA) isomerase, which is needed for phenanthrene and naphthanene degradation. We determined the nucleotide sequence of DNA fragment of 3271 bp which included the phnS gene. The fragment included an open reading frame of 594 bp which has ATG initiation codon and TAA termination codon and GGAA ribosomal binding site. The predicted amino acid sequence of the enzyme consists of 198 amino acids. The deduced amino acid sequence of the phnS enzyme exhibited 94％ identity with that of the corresponding enzyme in Sphingomonas aromaticivorans F199. The phnS gene is located downstream and in the same operon as phnQ and phnR, encoding a 2,3-dihydroxybiphenyl 1,2-dioxygenase and a ferredoxin component of biphenyl dioxygenase, respectively.

• Proceedings of the Microbiological Society of Korea Conference
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• 2002.10a
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• pp.175-179
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• 2002

The assemblies of our partial genomic sequence data of Sphingomonas chungbukensis DJ77, with the total size of 877,928 bp, was done by TIGR Assembler. The total size of our current obtained contigs was about 0.73 Mb. A comparative genome analysis between our uncompleted genome and the other completed genomes was performed by taking advantage of the availability of multiple complete genomes in COGs database (Clusters of Orthologous Groups of proteins) to produce the genomic prediction of our S. chungbukensis DJ77. This analysis based on homologues search among completed genomes provides good initial step to our better assigning putative function to predicted coding sequences.

• BMB Reports
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• v.42 no.3
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• pp.172-177
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• 2009

Phosphoglucose isomerase (PGI) is involved in synthesizing extracellular polysaccharide (EPS). The gene encoding PGI in Sphingomonas chungbukensis DJ77 was cloned and expressed in E. coli, and the protein was characterized. The pgi gene from DJ77 is 1,503 nucleotides long with 62% GC content and the deduced amino acid sequence shows strong homology with PGIs from other sources. The molecular masses of PGI subunit and native form were estimated to be 50 kDa and 97 kDa, respectively. Four potentially important residues (H361, R245, E330 and K472) were identified by homology modeling. The mutations, H361A, R245A, E330A, R245K and E330D resulted in decrease in Vmax by hundreds fold, however no significant change in Km was observed. These data suggest that the three residues (H361, R245Aand E330) are likely located in the active site and the size as well as the spatial position of side chains of R245 and E330 are crucial for catalysis.

• BMB Reports
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• v.42 no.8
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• pp.523-528
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• 2009

Phosphomannose isomerase (PMI) catalyzes the interconversion of fructose-6-phosphate and mannose-6-phosphate in the extracellular polysaccharide (EPS) synthesis pathway. The gene encoding PMI in Sphingomonas chungbukensis DJ77 was cloned and expressed in E. coli. The pmi gene is 1,410 nucleotides long and the deduced amino acid sequence shares high homology with other bifunctional proteins that possess both PMI and GDP-mannose pyrophosphorylase (GMP) activities. The sequence analysis of PMI revealed two domains with three conserved motifs: a GMP domain at the N-terminus and a PMI domain at the C-terminus. Enzyme assays using the PMI protein confirmed its bifunctional activity. Both activities required divalent metal ions such as $Co^{2+}$, $Ca^{2+}$, $Mg^{2+}$, $Ni^{2+}$ or $Zn^{2+}$. Of these ions, $Co^{2+}$ was found to be the most effective activator of PMI. GDP-D-mannose was found to inhibit the PMI activity, suggesting feedback regulation of this pathway.

• Microbiology and Biotechnology Letters
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• v.33 no.2
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• pp.84-89
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• 2005

Phosphomannomutase (PMM) is a key enzyme in prokaryotes and eukaryotes, which catalyzes the conversion of ${\alpha}$-D-mannose 6-phosphate to ${\alpha}$-D-mannose 1-phosphate. The latter is the substrate for the synthesis of GDP-mannose, which serves as the mannosyl donor for many metabolic pathways in the cells. We report here on the isolation of a gene from a genomic library of Sphingomonas chungbukensis DJ77, the pmmC gene encoding phosphomannomutase. The gene was cloned into E. coli expression vector, and the sequence was analyzed. The ribosomal binding site GGAAG lays 5 bp upstream of the ORF of 750 bp, which is initiated by ATG codon and terminated by TAG. The predicted sequence of the enzyme consists of 249 amino acids with a molecular mass of 27.4 kDa and showed $86.9\%$ similarity to that of eukaryotic phosphomannomutase after bioinformatical analyses with the conserved domain search of NCBI. The purified gene product revealed the activity of phosphomannomutase. In conclusion, we confirmed that pmmC gene encodes phosphomannomutase actually.

• Biomolecules & Therapeutics
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• v.16 no.2
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• pp.100-105
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• 2008

The sphingolipid metabolites act as lipid mediator for cell proliferation and apoptosis in mammalian cells. In bacteria, sphingolipid metabolism remains unknown. The purpose of this study was to investigate whether sphingolipid metabolism is potential target for fumonisin $B_1$($FB_1$) and desipramine in Sphingomonas chungbukensis, Gram-negative bacteria, by comparing the intracellular contents of bacterial sphingolipids with ones of HIT-T15 ${\beta}$-cells, hamster pancreatic cells. The concentrations of ceramide and dihydroceramide were 18.0 ${\pm}$ 12.0 and 0.025 ${\pm}$ 0.018 nmol/mg protein, respectively, in HIT-T15 cells. However, the concentrations of ceramide and dihydroceramide in the bacterial culture were 2.0 ${\pm}$ 1.2 and 10.6 ${\pm}$ 5.5 nmol/mg protein, respectively. $FB_1$ decreased the level of ceramide from 18.0 to 3.8 nmol/mg protein in HIT-T15 ${\beta}$-cells. However, dihydroceramide content in $FB_1$-treated HIT-T15 cells was slightly decreased compared with the control culture. When S. chungbukensis was treated with either $FB_1$ or desipramine, dihydroceramide level was increased by 5- and 4-fold, respectively, compared with the control bacteria. These results indicate that $FB_1$ and desipramine may act as an activator in bacterial sphingolipid biosynthetic pathway, and bacterial sphingolipid metabolism pathway appears to be different from the pathway of mammalian cells.