• Journal of Life Science
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• v.25 no.2
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• pp.121-129
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• 2015

Alginates are found in marine brown seaweeds and in extracellular biofilms secreted by some bacteria. Previously, we reported an oligoalginate lyase from Sphingomonas sp. MJ-3 (MJ3-Oal) that had an exolytic activity and protein sequence homology with endolytic polymannuronate (polyM) lyase in the N-terminal region. In this study, the MJ3-Oal was tested for both exolytic and endolytic activity by homology modeling using the crystal structure of Alg17c from Saccharophagus degradans 2-40T. The tyrosine residue at the $426^{th}$ position, which possibly formed a hydrogen bond with the substrate, was mutated to phenylalanine. The FPLC profiles showed that MJ3-Oal degraded alginate quickly to monomers as a final product through the oligmers, whereas the Tyr426Phe mutant showed only exolytic alginate lyase activity. $^1H$-NMR spectra also showed that MJ3-Oal degraded the endoglycosidic bond of polyM and polyMG (polymannuronate-guluronate) blocks. These results indicate that oligoalginate lyase from Sphingomonas sp. MJ-3 probably catalyzes the degradation of both exo- and endo-glycosidic bonds of alginate.

• Proceedings of the Korean Society of Life Science Conference
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• 2007.10a
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• pp.60.2-60.2
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• 2007

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• Korean Journal of Microbiology
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• v.33 no.2
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• pp.92-96
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• 1997

The 6.8 kb Xhol fragment of chromosomal ONA of Pseudomonas sp. 0177 contains the phnDEFG genes involved in the degradation of polyaromatic hydrocarbons and chlorinated aromatics. Here, we report the nucleotide sequence of the ORF encoding a polypeptide consisted of 143 amino acids with a Mr of 13,859. The nucleotide sequence of the ORF is 99% and 68.6% identical to the downstream region of catE of Sphingomonas sp. strain HV3 and the ORF between xylE and xylG of Sphingomonas yanoikuyae Bl, respectively. The deduced amino acid sequence of the PhnF has 62.3% identity with the amino acid encoded hy orfY region of Citrobacter freundii DSM30040. We now confirm that the ORF is located between the catechol 2,3-dioxygenase (C230), phnE, and 2-hydroxymuconic semialdehyde dehydrogenase (2HMSO), phnG.

• Journal of Microbiology and Biotechnology
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• v.31 no.3
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• pp.387-397
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• 2021

There is growing interest in the production of microalgae-based, high-value by-products as an emerging green biotechnology. However, a cultivation platform for Oocystis sp. has yet to be established. We therefore examined the effects of bacterial culture additions on the growth and production of valuable compounds of the microalgal strain Oocystis sp. KNUA044, isolated from a locally adapted region in Korea. The strain grew only in the presence of a clear supernatant of Sphingomonas sp. KNU100 culture solution and generated 28.57 mg/l/d of biomass productivity. Protein content (43.9 wt%) was approximately two-fold higher than carbohydrate content (29.4 wt%) and lipid content (13.9 wt%). Oocystis sp. KNUA044 produced the monosaccharide fucose (33 ㎍/mg and 0.94 mg/l/d), reported here for the first time. Fatty acid profiling showed high accumulation (over 60%) of polyunsaturated fatty acids (PUFAs) compared to saturated (29.4%) and monounsaturated fatty acids (9.9%) under the same culture conditions. Of these PUFAs, the algal strain produced the highest concentration of linolenic acid (C18:3 ω3; 40.2%) in the omega-3 family and generated eicosapentaenoic acid (C20:5 ω3; 6.0%), also known as EPA. Based on these results, we suggest that the application of Sphingomonas sp. KNU100 for strain-dependent cultivation of Oocystis sp. KNUA044 holds future promise as a bioprocess capable of increasing algal biomass and high-value bioactive by-products, including fucose and PUFAs such as linolenic acid and EPA.

• Journal of Microbiology
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• v.42 no.3
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• pp.243-247
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• 2004

Sphingomonas chlorophenolica ATCC39723 was successfully labeled with the gfp (green fluorescent protein) gene inserted into the pcpB gene by homologous recombination. As the gfp recombinant was easily distinguished from other indigenous organisms, the population of gfp recombinant was monitored after being released into the soil microcosms. Their population density dropped from 10$\^$8/ to 10$\^$6/ (cfu/$m\ell$) in the non-sterilized soil microcosms during the first 6 days. Moreover, the gfp recombinant was not detected even at lower dilution rates after a certain time period. The recombinant, however, survived for at least 28 days in the sterilized soil microcosms. Although the gfp recombinant did not degrade pentachlorophenol (PCP), this experiment showed the possibility of using gfp as a monitoring reporter system for S. chlorophenolica ATCC39723 and potentially other species of Sphingomonas.

• Journal of Microbiology and Biotechnology
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• v.13 no.6
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• pp.942-948
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• 2003

Soil samples were obtained from 5 sites contaminated with polycyclic aromatic hydrocarbons (PAHs). These soil samples were cultured in using phenanthrene as a sole carbon and energy source, and 36 strains of phenanthrene-degrading bacteria were isolated from 3 sites. Most of them degraded 500 ppm of phenanthrene within 8 to 10 days, and these isolates could degrade a few other PAHs other than phenanthrene. Their genotypes were determined by restriction digests of the l6S rRNA genes [amplified ribosomal DNA restriction analysis (ARDRA)]. It was found that all the phenanthrene degrading isolates were included in 4 ARDRA types, and they showed a strict site endemism. l6S rDNAs of 12 strains selected from different sites were sequenced, and they were all confirmed as Sphingomonas strains. Their l6S rDNA sequences were compared for phylogenetic analysis; their sequence showed a similar result to ARDRA typing, thus indicating that these heterotrophic soil bacteria are not regionally mixed. In addition, it was found that the microbial diversity among sampling sites could be monitored by l6S rDNA PCR-RFLP pattern alone, which is simpler and easier to perform, without l6S rDNA sequence analysis.

• Journal of Microbiology and Biotechnology
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• v.16 no.8
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• pp.1306-1310
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• 2006

Sphingomonas sp. strain SB5 could degrade carbofuran and carbofuran-7-phenol to a hydrolytic product, 2-hydroxy-3-(3-methlypropan-2-o1)phenol, and several red metabolites. However, the chemical structures of the red metabolites have largely remained unidentified. In this study, we identified the structure of one of the red metabolites as 5-(2-hydroxy-2-methyl-propyl)-2,2-dimethyl- 2,3-dihydro-naphtho[2,3-6]furan-4,6,7,9-tetrone by using mass spectrometric and NMR ($^1$H, $^{13}$C) analyses. It is suggested that the red metabolite resulted from condensation of some metabolites in the degradation of 2-hydroxy-3-(3-methlypropan-2-o1)phenol, a hydrolytic product derived from carbofuran. To our knowledge, this is the first paper to report a red metabolite in bacterial degradation of the insecticide carbofuran.

• Journal of Microbiology and Biotechnology
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• v.28 no.9
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• pp.1502-1510
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• 2018

Organic solvent-tolerant (OST) enzymes are widely applied in various industries for their activity and stability in organic solvents, for their higher substrate solubility, and for their greater stero-selectivity. However, the criteria for identifying OST enzymes largely remain undefined. In this study, we compared the amino acid composition of 19 OST esterases with that of 19 non OST esterases. OST esterases have increased the ratio of Ala and Arg residues and decreased the ratio of Asn, Ile, Tyr, Lys, and Phe residues. Based on our amino acid composition analysis, we cloned a carboxylesterase (EstSP2) from a psychrophilic bacterium, Sphingomonas glacialis PAMC 26605, and characterized its recombinant protein. EstSP2 is a substrate specific to p-nitrophenyl acetate and hydrolyzed aspirin, with optimal activity at $40^{\circ}C$; at $4^{\circ}C$, the activity is approximately 50% of its maximum. As expected, EstSP2 showed tolerance in up to 40% concentration of polar organic solvents, including dimethyl sulfoxide, methanol, and ethanol. The results of this study suggest that selecting OST esterases based on their amino acid composition could be a novel approach to identifying OST esterases produced from bacterial genomes.

• 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.