• Journal of Microbiology and Biotechnology
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• v.28 no.10
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• pp.1671-1682
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• 2018

Alginate lyases (endo and exo-lyases) are required for the degradation of alginate into its constituting monomers. Efficient bioethanol production and extraction of bioactives from brown algae requires intensive use of these enzymes. Nonetheless, there are few commercial alginate lyase preparations, and their costs make them unsuitable for large scale experiments. A recombinant expression protocol has been developed in this study for producing seven endo-lyases and three exo-lyases as soluble and highly active preparations. Saccharification of alginate using 21 different endo/exo-lyase combinations shows that there is complementary enzymatic activity between some of the endo/exo pairs. This is probably due to favorable matching of their substrate biases for the different glycosidic bonds in the alginate molecule. Therefore, selection of enzymes for the best saccharification results for a given biomass should be based on screens comprising both types of lyases. Additionally, different incubation temperatures, enzyme load ratios, and enzyme loading strategies were assessed using the best four enzyme combinations for treating Macrocystis pyrifera biomass. It was shown that $30^{\circ}C$ with a 1:3 endo/exo loading ratio was suitable for all four combinations. Moreover, simultaneous loading of endo-and exo-lyases at the beginning of the reaction allowed maximum alginate saccharification in half the time than when the exo-lyases were added sequentially.

• Journal of Life Science
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• v.17 no.5 s.85
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• pp.625-633
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• 2007

We isolated a new extracellular alginate lyase-producing microorganism, which displayed alginate-depolymerizing activity in plate assays, from coastal soils in Wando, Jeollanam-do, Korea. This alginate-depolymerizing bacterium belonged to the genus Streptomyces and it was named Streptomyces sp. MET 0515. An extracellular alginate lyase(ALY1) secreted by Streptomyces sp. MET 0515, was purified to homogeneity by a combination of acetone precipitation, anion-exchange chromatography (Q-Sepharose and DEAE-Sepharose) and Sephacryl S-200 HR gel filtration chromatography. Its molecular mass was 26 kDa as determined by SDS-PACE analysis. The enzyme had an optimal temperature of $70^{\circ}C$ for its activity, and was most active at pH 7.5. The thermal and pH stability were $0-50^{\circ}C$, and pH 6.0-9.0, respectively. The enzyme activity was stimulated by 1mM $Mn^{2+}$, and inhibited by 1mM $Fe^{3+}$, 1mM EDTA and 1mM $Zn^{2+}$. Preliminary analysis of substrate specificity showed that this alginate lyase had activity on both poly-alpha 1,4-L-guluronate and poly-beta 1,4-D-mannuronate in the alginate molecule.

• Proceedings of the Korean Society of Life Science Conference
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• 2006.09a
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• pp.86.1-86.1
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• 2006

• Proceedings of the Korean Society of Life Science Conference
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• 2006.09a
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• pp.85.2-85.2
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• 2006

• Food Engineering Progress
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• v.21 no.2
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• pp.103-109
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• 2017

Alginate lyase from Streptomyces violaceoruber was purified by DEAE sephacel chromatography and SP sepharose chromatography. The specific activity of the purified enzyme was 14.6 units/mg protein, representing a 40.6-fold purification of the crude extract. The final preparation thus obtained showed a single band on Tricine-SDS polyacrylamide gel electrophoresis whose molecular weight was determined to be 23.3 kDa. The polyMG block of sodium alginate was hydrolyzed by the purified alginate lyase and then separated by activated carbon column chromatography and bio gel P-2 gel filtration. The main hydrolysates were composed of hetero type M/G-oligosaccharides with the degrees of polymerization (D.P.) being 6 and 8. To investigate the effects of hetero type M/G-oligosaccharides from the sodium alginate on the growth of some intestinal bacteria, cells were cultivated individually on the modified-MRS medium containing D.P. 6 and 8 M/G-oligosaccharides. B. longumgrew 4.25-fold and 6.44-fold more effectively by the treatment of D.P. 6 and 8 M/G-oligosaccharides compared with those of standard MRS medium. In addition, B. bifidumgrew 3.3-fold and 5.4-fold more effectively by the treatment of D.P. 6 and 8 M/G-oligosaccharides. In conclusion, D.P. 8 was more effective than D.P. 6 hetero M/G-oligosaccharides as regards the growth of Bifidobacteriumspp. and Lactobacillus spp.

• Korean Journal of Microbiology
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• v.54 no.4
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• pp.463-464
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• 2018

Tamlana sp. UJ94 isolated from seawater can degrade alginate. To identify the genomic basis of this activity, the genome was sequenced. The genome was composed of 4,116,543 bp, 3,609 coding sequences, and 35.2 mol% G + C content. A BLASTp search predicted the presence of 9 alginate lyases as well as 6 agarases, 5 amylases, 4 carrageenases, 1 cellulase, 4 pectate lyases, and 7 xylanases, indicating its ability to degrade diverse polysaccharides. The genome of strain UJ94 is a source of polysaccharide-degrading enzymes for bioconversion processes.

• Applied Chemistry for Engineering
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• v.23 no.2
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• pp.164-168
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• 2012

We investigated for the production of biological bio-ethanol from Laminaria japonica using the hydrolysis reaction of enzymes and organic acids and the polysaccharide content was also analyzed. The composition of the polysaccharide was characterized as 65.99% alginate, 6.24% laminaran and 27.77% mannitol. The optimum concentration for reducing the sugar conversion by Laminaria japonica was found to be 1.874 g/L at an acetic acid concentration of 1.5%, $121^{\circ}C$ for 60 min, and for an ascorbic acid of 2.0%, 4.291 g/L was produced in the same condition. The enzyme hydrolysis such as alginate lyase and laminarinase contained the maximum 2.219 g/L reducing sugar. In the result of ethanol fermentation using hydrolysate of Laminaria japonica, the organic acid treatment showed a high of reducing sugar yield, but decreased the ethanol yield, and then the maximum ethanol production obtained was 1.26 g/L using the mixed treated of enzyme.

• Journal of Microbiology and Biotechnology
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• v.23 no.6
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• pp.872-877
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• 2013

In a previous study, we isolated and reported a second species of the Saccharophagus genus, Saccharophagus sp. strain Myt-1. In the present study, an alginate lyase gene (algMytC) from the genomic DNA of Myt-1 was cloned and characterized. The DNA sequence fragment obtained contained an open reading frame of 1,032 bp that encoded a protein of 343 amino acids with an estimated molecular mass of 37.6 kDa and a pI of 6.60. The deduced protein, AlgMytC, had the conserved amino acid sequences (RTELREM, QIH, YFKAGVYNQ) of the polysaccharide lyase family 7. A BLAST homology search indicated that AlgMytC shared an amino acid sequence identity of 95.9% with alg7A of S. degradans 2-40. The cloned and purified AlgMytC protein showed optimal activity at $40^{\circ}C$, and retained more than 90% of its total activity even after treatment at $25^{\circ}C$ for 24 h. AlgMytC was very alkaliphilic with an optimal pH of 9.0, and more than 90% of its activity was retained in the pH range 8.5-10.0. Moreover, AlgMytC was stable over a wide pH range. The activity of AlgMytC was also stable in the presence of various detergents.

• Molecular & Cellular Toxicology
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• v.4 no.4
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• pp.307-310
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• 2008

AAD16034 is an alginate lyase from Pseudoalteromonas sp. IAM14594. A very close homologue with known 3D structure exists (marine bacterium Pseudoalteromonas sp. strain no. 272). A three-dimensional structure of AAD16034 was generated based on this template (PDB code: 1J1T) by comparative modeling. The modeled enzyme exhibited a jelly-roll like structure very similar to its template structure. Both enzymes possess the characteristic alginate sequence YFKhG+Y-Q. Since AAD16034 displays enzymatic activity for poly-M alginate, docking of a tri-mannuronate into the modeled structure was performed. Two separate and adjacent binding sites were found. The ligand was accommodated inside each binding site. By considering both binding sites, a plausible binding pose for the poly-M alginate polymer could be deduced. From the modeled docking pose (i.e., the most important factor that attracts alginate polymer into this lyase) the most likely interaction was electrostatic. In accordance with a previous report, the hydroxyl group of Y345 was positioned close to the ${\alpha}$-hydrogen of ${\beta}$-mannuronate, which was suitable to initiate a ${\beta}$-elimination reaction. K347 was also very near to the carboxylatemoiety of the ligand, which might stabilize the dianion intermediate during the ${\beta}$-elimination reaction. This implies that the characteristic alginate sequence is absolutely crucial for the catalysis. These results may be exploited in the design of novel enzymes with desired properties.

• Journal of Microbiology and Biotechnology
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• v.11 no.1
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• pp.118-123
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• 2001

The alginate lyase A1-III gene of Sphingomonas species A1 is composed of 1,077 nucleotides, encoding a protein (359 amino acids) with a molecular mass of 40,322 Da. Recombinant A1-III expressed in Escherichia coli exhibited the same full enzymatic activity as native A1-III. In order to identify the critical residue for activity, a site-directed mutation was introduced into the A1-III gene (H192A, His192->Ala). Recombinant A1-III (H192A) exhibited a significant decrease in enzyme activity (one-thirty thousandth of that of A1-III), without any conformational change, as detected by the CD spectra in the far UV region. Also, the chemical modification of wild-type A1-III with methyl 4-nitro benzene sulfonate resulted in a 40% decrease from the initial activity, whereas the same modification of A1-III (H192A) produced no change in the activity. The role of His192 on the catalytic process was also explored based on a model of A1-III docked with mannuronic acid into the active site.