• Proceedings of the Korean Society for Bioinformatics Conference
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• 2003.10a
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• pp.26-51
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• 2003

Large scale protein interaction maps provide a new, global perspective with which to analyse protein function. PSIMAP, the Protein Structural Interactome Map, is a database of all the structurally observed interactions between superfamilies of protein domains with known three-dimensional structure in thePDB. PSIMAP incorporates both functional and evolutionary information into a single network. It makes it possible to age protein domains in terms of taxonomic diversity, interaction and function. One consequence of it is to predict the most important protein domain structure in evolution. We present a global analysis of PSIMAP using several distinct network measures relating to centrality, interactivity, fault-tolerance, and taxonomic diversity. We found the following results: ${\bullet}$ Centrality: we show that the center and barycenter of PSIMAP do not coincide, and that the superfamilies forming the barycenter relate to very general functions, while those constituting the center relate to enzymatic activity. ${\bullet}$ Interactivity: we identify the P-loop and immunoglobulin superfamilies as the most highly interactive. We successfully use connectivity and cluster index, which characterise the connectivity of a superfamily's neighbourhood, to discover superfamilies of complex I and II. This is particularly significant as the structure of complex I is not yet solved. ${\bullet}$ Taxonomic diversity: we found that highly interactive superfamilies are in general taxonomically very diverse and are thus amongst the oldest. This led to the prediction of the oldest and most important protein domain in evolution of lift. ${\bullet}$ Fault-tolerance: we found that the network is very robust as for the majority of superfamilies removal from the network will not break up the network. Overall, we can single out the P-loop containing nucleotide triphosphate hydrolases superfamily as it is the most highly connected and has the highest taxonomic diversity. In addition, this superfamily has the highest interaction rank, is the barycenter of the network (it has the shortest average path to every other superfamily in the network), and is an articulation vertex, whose removal will disconnect the network. More generally, we conclude that the graph-theoretic and taxonomic analysis of PSIMAP is an important step towards the understanding of protein function and could be an important tool for tracing the evolution of life at the molecular level.

• Microbiology and Biotechnology Letters
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• v.44 no.4
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• pp.504-511
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• 2016

Lactulose, a synthetic disaccharide, has received increasing interest because of its role as a prebiotic that can increase the proliferation of Bifidobacterium and Lactobacillus spp. and enhance the absorption of calcium and magnesium. While the industrial production of lactulose is still mainly achieved by the chemical isomerization of lactose in alkaline media, this process has drawbacks including the need to remove catalysts and by-products, as well as high energy requirements. Recently, the use of cellobiose 2-epimerase (CE) has been considered an interesting alternative for industrial lactulose production. In this study, to develop a process for enzymatic lactulose production using CE, we screened improved mutant enzymes ($CS-H^RC^E$) from a library generated by an error-prone PCR technique. The thermostability of one mutant was enhanced, conferring stability up to $75^{\circ}C$, and its lactulose conversion yield was increased by 1.3-fold compared with that of wild-type CE. Using a recombinant Escherichia coli strain harboring a CS35 $H^RC^E$-expressing plasmid, we prepared cell beads immobilized on a Ca-alginate substrate and optimized their reaction conditions. In a batch reaction with 200 g/l lactose solution and the immobilized cell beads, lactose was converted into lactulose with a conversion yield of 43% in 2 h. In a repeated 38-plex batch reaction, the immobilized cell beads were relatively stable, and 80% of the original enzyme activity was retained after 4 cycles. In conclusion, we developed a reasonable method for lactulose production by immobilizing cells expressing thermostable CE. Further development is required to apply this approach at an industrial scale.

• Applied Biological Chemistry
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• v.37 no.6
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• pp.451-455
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• 1994

To detect naturally occurring bioactive compounds from unused marine resources, the screening for the 5-lipoxygenase(potato lipoxygenase-II) inhibitors in Asterina pectinifera, Halocynthia roretzi skin, Nototodarus sloani ink, Anthocidaris crassispina skin, Sargassum horneri, Agarum cribrosum, Odonthalia corymbifera and Desmarestia ligulata was carried out. Water, ether, acetone and methanol fractions extracted from Sargassum horneri had strong inhibitory effect on enzymatic lipid oxidation by potato lipoxygenase-II, and their $IC_{50}$ were 320, 18, 9.5 and $100\;{\mu}g/mL$, respectively. The $IC_{50}$ of ether fraction extracted from Asterina pectinifera and acetone fraction extracted from Nototodarus sloani ink were 29.5 and $34.3\;{\mu}g/mL$, and these extracts showed relatively excellent inhibitory activity. Nonpolar solvent (ether, acetone) extracts of tested marine organisms had more inhibitory effect against 5-lipoxygenase than the polar solvent(water) extracts.

• Journal of Microbiology and Biotechnology
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• v.24 no.6
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• pp.748-755
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• 2014

In the genome annotation of Escherichia coli MG1655, the orf382 (1,149 bp) is designated as a gene encoding an alcohol dehydrogenase that may be Fe-dependent. In this study, the gene was amplified from the genome by PCR and overexpressed in Escherichia coli BL21(DE3). The recombinant $6{\times}$His-tag protein was then purified and characterized. In an enzymatic assay using different hydroxyl-containing substrates (n-butanol, $\small{L}$-threonine, ethanol, isopropanol, glucose, glycerol, $\small{L}$-serine, lactic acid, citric acid, methanol, or $\small{D}$-threonine), the enzyme showed the highest activity on $\small{L}$-threonine. Characterization of the mutant constructed using gene knockout of the orf382 also implied the function of the enzyme in the metabolism of $\small{L}$-threonine into glycine. Considering the presence of tested substrates in living E. coli cel ls and previous literature, we believed that the suitable nomenclature for the enzyme should be an $\small{L}$-threonine dehydrogenase (LTDH). When using $\small{L}$-threonine as the substrate, the enzyme exhibited the best catalytic performance at $39^{\circ}C$ and pH 9.8 with $NAD^+$ as the cofactor. The determination of the Km values towards $\small{L}$-threonine (Km = $11.29{\mu}M$), ethanol ($222.5{\mu}M$), and n-butanol ($8.02{\mu}M$) also confirmed the enzyme as an LTDH. Furthermore, the LTDH was shown to be an ion-containing protein based on inductively coupled plasma-atomic emission spectrometry with an isoelectronic point of pH 5.4. Moreover, a circular dichroism analysis revealed that the metal ion was structurally and enzymatically essential, as its deprivation remarkably changed the ${\alpha}$-helix percentage (from 12.6% to 6.3%).

• Microbiology and Biotechnology Letters
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• v.31 no.2
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• pp.140-144
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• 2003

Type Ⅰ signal peptidase is an integral membrane protein that functions to cleave signal peptides from secreted and membrane proteins. The enzyme serves as a potential target for the development of novel antibacterial agents due to its unique physiological properties. Despite being one of the best characterized enzymes, the catalysis of Type Ⅰ signal peptidase still remains controversy over the catalytic serine/lysine dyad mechanism. It appears that the dyad proteases are generally less efficient than the prototypical serine/histidine/aspartic acid triad found in most enzymes, although Type Ⅰ signal peptidase is an exception to this rule. In this paper, we have proposed that Type Ⅰ signal peptidase may act as the serine/lysine/aspartic acid triad cataltytic mechanism. Therefore, the aspartic acid 99 residue in the E. coli signal peptidase was chosen and mutated to an alanine to see if there is any possible role of the aspartic acid in the catalytic function. Type Ⅰ signal peptidase D99A protein was inactive in vitro assay using the procoat synthesized by in vitro transcription translation. However, the mutant was active using a highly sensitive in vivo assay. Pulse-chase experiments show that the replacement of aspartic acid 99 with alanine results in a very unstable signal peptidase molecule. Therefore, we conclude that it is unlikely that the residue is directly involved in catalysis, but rather plays an important role in stabilizing the protein structure.

• Nutrition Research and Practice
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• v.1 no.4
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• pp.371-375
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• 2007

To control blood glucose level as close to normal is a major goal of treatment of diabetes mellitus. Hyperglycemia and hyperlipidemia are the major risk factors for cardiovascular complications, the major cause of immature death among the patients with type 2 diabetes. The purpose of this study is to determine the hypoglycemic and hypolipidemic effects of Salicornia herbacea in animal model of type 2 diabetes and to investigate the possible mechanisms for the beneficial effects of S. herbacea. S. herbacea was extracted with 70% ethanol and desalted with 100% ethanol. Three week-old db/db mice (C57BL/KsJ, n=16) were fed AIN-93G semipurified diet or diet containing 1% desalted ethanol extract of S. herbacea for 6 weeks after 1 week of adaptation. Fasting plasma glucose, triglyceride, and total cholesterol were measured by enzymatic methods and blood glycated hemoglobin ($HbA_{1C}$) by the chromatographic method. Body weight and food intake of S. herbacea group were not significantly different from those of the control group. Fasting plasma glucose and blood glycated hemoglobin levels tended to be lowered by S. herbacea treatment. Consumption of S. herbacea extract significantly decreased plasma triglyceride and cholesterol levels (p<0.05). The inhibition of S. herbacea extract against yeast ${\alpha}$-glucosidase was 31.9% of that of acarbose at the concentration of 0.5 mg/mL in vitro. The inhibitory activity of ethanol extract of S. herbacea against porcine pancreatic lipase was 59.0% of that of orlistat at the concentration of 0.25 mg/mL in vitro. Thus, these results suggest that S. herbacea could be effective in controlling hyperlipidemia by inhibition of pancreatic lipase in animal model of type 2 diabetes.

• Polymer(Korea)
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• v.30 no.3
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• pp.259-265
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• 2006

Small intestinal submucosa (SIS) consists of some growth factors which can stimulate cell activity, and PVA has been widely utilized in the area of wound dressing as hydrogel which is easy to be removed from wounds. In this study, native SIS sheets were coated with PVA by immersing them into 2, 4, and 10 wt% of PVA solution and then lyophilized on two type of molds to endow the prepared wound dressing with easy removal property from wounds. The mechanical properties were examined through tensile test. Moreover, enzymatic degradation, water uptake, and in vitro test were carried out to characterize the prepared SIS sheets. The tensile strength of the SIS sheets coated with PVA (PVA-SIS) were decreased, whereas the elongation were increased. Degradation ratio of the PVA-SIS sheets was decreased compare to native SIS. Water uptake ability was improved at 2 and 4 wt% of PVA. The degree of fibroblast attachment was lower than the native SIS sheets. In conclusion, this study suggests that the PVA coated SIS sheets have a potential for the applications of wound dressing and biodegradable injectable materials.

• KSBB Journal
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• v.21 no.6 s.101
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• pp.474-478
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• 2006

For the economically feasible production of ethanol, utilization of SFW (saccharified food wastes) as substrate for synchronous saccharification and fermentation (SSF) process was developed in this study. When 200 g of food wastes and 40 mL of enzyme ($amylase activity,\;3.0\;U/m{\ell}$) were reacted, production rate of reducing sugar was $5.84\;g/{\ell}{\cdot}h$, and consumption rate was $-3.88\;g/{\ell}{\cdot}h\;at\;35^{\circ}C$ So suitable condition of SSF was concluded at temperature of $35^{\circ}C$. Also, optimal enzyme concentration of SSF was concluded in $2.0\;U/m{\ell}$, at this condition, the production rate of reducing sugar was $4.80\;g/{\ell}{\cdot}h$ At SSF process, when 50 g of food wastes was supplied in 12 h interval, $64\;g/{\ell}$ of ethanol and 0.45 g-ethanol/g-reducing sugar in yield were obtained in 120 h fermentation. Thus, the technology of high yield of ethanol production using food wastes was confirmed. And semi-continuos SSF system for cutting off cost of enzymatic saccharification was developed in this study.

• Journal of the Korean Society of Food Science and Nutrition
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• v.36 no.12
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• pp.1596-1603
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• 2007

This study was conducted to improve functional properties of salmon frame extracts using various commercial enzymes (Alkalase 2.4 L FG, Flavourzyme 500 MG, Neutrase 0.8 L and Protamex 1.5 MG). The ACE (angiotensin I converting enzyme) inhibitory activity was the highest ($IC_{50}=0.67mg/mL$) in the product incubated with Neutrase for 4 hrs (N4-treated hydrolysates) among the various extracts incubated with commercial enzymes for different times. However, antioxidant activities of all salmon frame extracts were less than 15%. There were no significant differences in the proximate composition and sensory evaluation of the fish odor and taste. However, N4-treated hydrolysate was improved in the extractive-nitrogen content and transmission compared to the other enzymatic hydrolysates. When compared to commercial Gomtang products, N4-treated hydrolysate was also high in protein, extractive-nitrogen, total amino acid, and calcium contents, while low in taste sensory score. There were no differences in transmission and sensory score on the fish odor between N4-treated hydrolysates and commercial Gomtang.

• Korean Journal of Food Science and Technology
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• v.25 no.1
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• pp.39-45
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• 1993

The effects of enzymatic modification with pepsin and actinidin was studied on molecular weight distributions and functional properties of hydrolysates from soy protein isolate (SPI) differing in degree of hydrolysis. The hydrolyzed SPI by pepsin showed 41.5% degree of hydrolysis after 5 min, and maximum hydrolysis was obtained after 2 hours. Actinidin hydrolyzed SPI 26.71% degree after 1 hour. On SDS-PAGE, native SPI showed 9 distinguishable bands on SDS-PAGE gel. Pepsin treated SPI showed one broad band in the lower part of gel. This band was shifted further to the bottom of the gel and became faint as hydrolysis time increased. While actinidin treated SPI showed different SDS-PAGE pattern from pepsin. However PAGE patterns were similar with pepsin and actinidin treated groups. With pepsin treatment, solubility of SPI distinctively increased around isoelectric point(pI). Emulsifying activity (EA) and emulsifying stability (ES) showed marked increase over pH range of $3.0{\sim}8.0$. 5 min modified group had most excellent foam expansion (FE). Foam stability (FS) was increased as pepsin treatment time increased at pI. With actinidin treatment, solubility was increased. 60 min modified SPI had the most effective EA at pH 4.5. However ES was not effected by actinidin treatment. 5 min modified group was most effect in FE. FS was higher at alkaline pH.