Earticle

Palladium is one of the most expensive metals which can catalyze many chemical reactions. Recovery of precious metals from industrial wastes is very difficult and consumes a lot of time and money. In spite of these disadvantages it would be still profitable if these metals are recovered. This work estimated sorption characteristics of Pd(II) ion by Corynebacterium glutamicum and recovered it from the Pd(II)-sorbed biomass. Palladium biosorption performed well at pH 3. Sorption isotherm at pH 3 was determined to be 137.6 mg/g by the Langmuir model. Kinetic experiments revealed that the sorption equilibrium was attained within 1 h, which was much faster than that of many reported sorbents. After sorption, the Pd(II)-loaded biomass was charred at 600 oC for 3 h, and the ash bound with metal ions was dissolved in aqua regia for metal recovery. Based on the ICP analysis, the recovery rate of Pd(II) ions was calculated as a relative value from the initial amount of the bound metal ions. From the above results it could be concluded that C. glutamicum biomass would be a useful raw material for low-cost and efficient biosorption and recovery of precious metals.

In the summer, to avoid propagation of green algae bloom. Copper sulfate was scattered on reservoir. The copper leads to damage fluidity and permeability of the membrane by combining with cell membrane temporarily. Nutrient salts eluted by the sediment of polluted reservoir, as important nutrient source of algae, caused sudden breeding of green algae, and which also exacerbate reservoir by generating toxic materials.(1) It was changed the concentration of Copper(Ⅱ) sulfate pentahydrate in 0~3 μg/ml using Scenedesumus. sp. of the fresh water algae in this study and measured a cell number, OD, a change of the time of the chlorophyl content, and LC50(72hr). As a result, LC50 was calculated 2.35 μg/ml and predicted no effects concentration (PNEC) of 23.5 ng/ml is derived and considers fact water system environment for applying uncertainty calculation 100. It was detected in 0.051~0.167 mg/l as for the copper of the cavity water of the reservoir (Dongbok) sedimentation layer2) and detected in 0.021~0.110 mg/l (2007) as for the copper concentration of reservoir (Dongbok) outer layer. The toxicity was predicted based on experimantal results, the copper toxocity could influenced to benthos of sedimental layer and outer layer organism, but the stability evaluation for the fact water ecosystem must be examined more concretely.

Bioethanol is in the limelight as the ideal alternative fuel. It is one of the most clean and sustainable energy resources in the near future. This fuel can be classified into starchy and cellulosic ethanol according to the starting materials. Cellulosic ethanol is on investigation because of its great amount of mass. Cellulose has substantial stability and low conversion efficiency to glucose that can be further fermented to alcohol. For decades, scientists looked for the enhancement of activity of cellulase system by gene manipulation. However, there were few studies of the mixed culture for bioethanol production. Trichoderma reesei is one of the most well-studied fungi producing cellulase. Although Trichoderma reesei yields high activity of endoglucanase and exoglucanase, its drawback is the low activity of ß-glucosidase of the cellulase system. Effective Microorganisms(EM) are in the combination of various microorganisms consisting mainly of lactic acid bacteria, purple bacteria, yeast, actinomycetes, and fungi. Because Aspergillus oryzae and Lactobacillus plantarum in EM can produce ß-glucosidase, they can be compatible with T. reesei to increase the hydrolysis of cellulose.

This work focused on making a comparative study on the biosorbents and commercial sorbents for the recovery of nickel-containing effluents. The variety of chemically modified the waste Escherichia coli biomass from a fermentation industry as the biosorbents. Raw (RB), citric acid-treated (CAB), phosphoric acid-treated (PAB) and succinated (SB) biomass were evaluated as sorbents to remove Ni(II) ions from aqueous solutions. The results were compared with those from using Amberlite IRN-150 and IR-120 resin as commercial sorbent resins. All sorbents performed well in Ni(II) removal at pH > 5, but considering the nickel speciation and precipitation, the biosorption performance was optimized at pH 6. The modified biomass (SB (39.11 mg/g), CAB (47.14 mg/g) and PAB (46.24 mg/g)) showed the higher sorption capacity than raw biomass (RB (31.31 mg/g)). The Amberlite IR-120 resin showed the highest sorption capacity (68.85 mg/g). But, desorption efficiency of nickel ions from Amberlite IRN-150 and IR-120 resin were 20% below, which was too low for the sorption performance of the resin to be maintained at an economic level in subsequent cycles. In contrast, biomass was 70% over. The capability for recovery of the metal ions is the main advantages of the bacterial waste biomass, in addition to its low cost. It deserves further investigations into the details of practical application, for example, on the development of surface modified, sorption process optimization and desorption methods.

The effect of the phospholipase A2-treatment of bacterial biomass on the biosorption of precious metals was studied. The pre-treatment by phospholipase A2 confirmed the important role of carboxylic groups in the binding of the metals. The pre-treatment of the bacterial biomass with a phospholipase A2 led to a significant increase of sorption capacity. Actually, an increase of 50-70% of the sorption capacity was obtained when the pre-treatment was carried out for 5 h with a solution containing 0.3% of phospholipase solution at 37 0C. Lysozyme/papain-treatment before phospholipase A2-treatment was carried out to make easier the phospholipase penetrate through the peptidoglycan layer.

Xylanase (EC 3.2.1.8) is the name given to a class of enzymes which degrade the linear polysaccharide beta-1,4-xylan into xylose, thus breaking down hemicellulose, which is a major component of the cell wall of plants. A new xylanase gene, KRICT PX1 isolated from Paenibacillus sp. HPL-001 was expressed in E. coli, and the biochemical properties of the enzyme was examined. The GST-fused xylanase was purified with immobilized glutathione column, and xylanase activity of the purified GST-fused protein was 10.97 U/mg protein with Km value of 3.16 for xylan from birch wood. The optimum pH and temperature for the activity of the enzyme were 6~7 and 40~50°C, respectively. Most salts, such as NaCl, LiCl, KCl, NH4Cl, CaCl2, MgCl2, MnCl2, and CsCl2 did not change the enzyme activity at 1 mM except CuSO4, ZnSO4, and FeCl3. And also, 1 mM of EDTA, 2-mercaptoethanol, and PMSF were not effective on the enzyme activity. The enzymatic products resulting from the reaction with xylan from birch wood was mostly xylose with a small amount of arabinose.

Cellulose is the most renewable resource in the world. It is a linear polymer with consists of repeated units of monomer glucose with β-1,4-glycosidic bond. This highly crystallined structure of cellulose makes the reaction slow in cellulase-catalyzed hydrolysis reaction in aqueous media because of the inaccessibility of enzyme and water to their adsorption sites on cellulose. Therefore, the pretreatment of cellulose in order to increase the surface area which provides easy access of water and cellulase, is required1-3. This results in improvements in hydrolysis kinetics and conversion of cellulose to glucose. Rate of hydrolysis in cellulose-dissolving Ionic Liquid, such as 1-butyl-3-methylimidazolium chloride ([Bmim][Cl]) was improved by approximately 50-fold higher for regenerated cellulose than the rate of untreated cellulose(Avicel PH-101)4. In this study, influence of treatment with [Bmim][Cl] that do not dissolve cellulose on initial rate of enzymatic hydrolysis was compared with that of Ionic Liquids those dissolve cellulose.

The Bioethanol for use as a liquid fuel by fermentation of renewable biomass as an alternative to petroleum is important from the viewpoint of global environmental protection. After fermentation, also Dried Distiller Grain with Soluble(DDGS) which is very useful as feed is obtained as a byproduct. The objective of this work is to evaluate ethanol productivity of Gangdaok(domestic corn) and DDGS using low-temperature pretreatment cost efficient process compared to a pressure pretreatment process. First, conditions are optimized for low-temperature pretreatment process. It was found that at 98℃ the reaction time was 1.5h. According to the fermentation results, it shows almost same ethanol productivity as the low-temperature pretreatment process and the pressure pretreatment process. Also, as a result of DDGS, Gangdaok has more essential amino acid (21.1mg/g) than that of Ambtosia (USA corn), which is a widely used materials as feedstock. To conclude, it means the one excels in economy and value of feedstock than the others.

The objectives of this study were to develope the economical process for bioethanol production from domestic triticale and investigate optimal fermentation conditions such as temperature, time, and enzyme concentration used to pre-treatment process. Triticale mash, containing 148 g of total sugar per 1 L of mash, was fermented with Saccharomyces cerevisiae CHY1011 at 33℃. Fermentation of mash supplemented with enzyme was completed within 48 - 60 hours, and the ethanol yield was 410.9 L/tonne of dry base. On the other hand, fermentation of mash without enzyme addition was completed within 36 - 48 hours, but the ethanol yield was 342.2 L/tonne of dry base. For optimal bioethanol production from triticale, viscosity reduction enzyme was added in the pre-treatment process, and the fermentation rate of triticale was 92.0-94.2%. In addition, the results showed that bioethanol production of triticale by low-temperature pre-treatment would provide higher ethanol production efficiency and lower operating costs.

Freundlich isothermal adsorption parameters, applicable to such biofilter-model as process-lumping model(Lim's model), for sterilized granular activated carbon(GAC), sterilized compost and sterilized equal volume mixture of GAC and compost were obtained and were compared each other. Adsorption capacity parameters(K) and adsorption exponents of Freundlich adsorption isotherm equation, which simulates the adsorbed amount of ethanol equilibrated with the ethanol concentration of the condensed water in the pore of the media, were constructed for sterilized granular activated carbon(GAC), sterilized compost and sterilized equal volume mixture of GAC and compost as (0.7566 and 5.070x10-7), (0.8827 and 1.000x10-8) and (0.5688 and 5.243x10-6), respectively. These Freundlich isothermal adsorption parameters were applicable to the adsorption characteristics of biofilter media enclosed with bio-layer. The order of magnitude of the ratio of ethanol-air/water partition coefficient and toluene-air/water partition coefficient was almost consistent to that of ethanol-adsorbed amounts in this experiment with compost and in the investigation of Delhomenie et al. on toluene-adsorption to wet compost.

Experiment and its mathematical modeling were preformed in order to predict the operation of a reactor combined of an aerobic and an anaerobic reactor to remove ammonia from waste-air stream. The aerobic reactor is identical to a three phase fluidized-bed biofilm reactor whereas the anaerobic one is a two phase fluidized-bed biofilm reactor. The model was built based upon some basic processes of chemical engineering and biotechnology such as mass transfer and biochemical reaction. According to the experimental results it has been shown that a high removal efficiency of ammonia can be achieved higher than 90% at early state of the reactor-performance. However it was found that this value decreases and maintains at ca. 80% when it reaches the steady state of operation. With slight difference less than 12% between model-prediction and the experimental data, the results of this study have shown the relevant correlation between them

Recent reports describe intensively the technical and economical views of the bioethanol production. Particularly, there have been many interests in the use of lignocellulosic material from the agricultural by-product for the resource of the bioethanol production, because of the price rises in crops (e.g. corn and rice etc.). Among those, sugar cane bagasses, sorghum straw, wheat straw, rice straw, corn stover, and corncob are being investigated on the usage as a resources of bioethanol production, in which the strategy for their hydrolyses, their contents of fermentable sugar, and the fermentation process were examined. Prior to the bioethanol fermentation, the hydrolyses of those by-products were popularly carried out using the dilute acid solutions of sulfuric acid, phosphoric acid, and hydrochloric acid. In addition, there has been reported the enzymatic hydrolysis of lignocellulosic by-product, and the combined hydrolysis protocol using enzyme and dilute acid was also attempted. Particularly the corncob is not only an agricultural by-product, but also a residue produced from the corn processing industry. While, in occidental countries, the corn stover has been considered as an alternative resource for bioethanol production, the corncob is examined in Far-east Asian countries. Hydrolysate of conrcob contains 60-80% of glucose, xylose, and arabinose in dry basis. In this study, we investigated on the hydrolysis of corn hull, by-product of corn starch industry, and its feasibility of bioethanol production, in which Pachysolen tannophilus was used as C6 and C5 sugar-utilizing yeast. Particularly the optimized concentration of acid, and the amount of corn hull for hydrolysis were examined. The pretreatment method for hydrolysate was also investigated.

Food wastes, dumped from the Seoul metropolitan area, were used for the production of biohydrogen. A food waste product (FWP), homogenized and dried, was obtained from a company, E-Service Corp., and used as growth media and carbon sources for an anaerobic acidophilic Clostridium sp. Clostridium acidisoli KCTC 5384 produced H2 using the FWP regardless of the additional chemicals or carbon sources for growth of the bacteria under anaerobic culture condition. In a medium containing 12% (w/v) FWP, C. acidisoli KCTC 5384 produced the maximum hydrogen production potential of 128 ml H2 l-1h-1. Hydrogen production rate was changed according to the amount of FWP in medium. Increase of the amount of FWP decreased the hydrogen production rate.

The operating conditions for a hollow-fiber membrane filter reactor containing methanogens that can utilize carbon dioxide and hydrogen was studied for the reduction of carbon dioxide. Anaerobic sludge was inoculated into each reactor (20% of the working volume). To determine the optimum conditions for methane production, two reactors (CRS1, CRS2) were operated for 100 days under different pH conditions, i.e., pH 5 and pH 7, with both reactors under an input gas pressure of 2～3 psi and 37 ℃. Methane was produced in the acidic reactor after 2 days and the methane percentage in the biogases produced was 80% on day 5. The volume of methane produced was on average 200～300 ml/day. The neutral reactor, CRS2, however, saw methane produced after only 2 hours. The methane in the biogas increased to 60% with nearly 2-times more than the acidic reactor, i.e., 400～600 ml/day on average over the test period.

The biodiesel industry has increased due to its eco-friendly and renewable alternative biodiesel. Because 10 kg of biodiesel yields 1 kg of glycerol, the cost of crude glycerol is rapidly decreased in the world market. There is a great advantage to use crude glycerol for the useful chemical productions from economical and environmental standpoints. In this work, Clostridium pasteurianum DSM 525, C. butyricum DSM 2477 and C. diolis DSM 15410 were tested for the production of butanol and 1,3-propanediol (1,3-PD) with crude glycerol. Because crude glycerol without pre-purification exhibited the inhibitory effect on the growth of the tested Clostridium species, crude glycerol was treated with 5 N HCl to remove soap components which are known as cell growth inhibitors. The color of crude glycerol was changed from dark brown to light yellow after pretreatment. The growth of microorganisms was enhanced with the pretreated crude glycerol and 4~5 g/L of 1,3-PD and 5~6 g/L of butanol were produced in the pretreated crude, which were similar to those produced in pure glycerol (5 g/L). In conclusion, the production of 1,3-PD and butanol with clostridium species can be achieved with crude glycerol after the simple pretreatment.

Biobutanol, one of the promising biofuels for future, has the potential to meet the needs of sustainable and green energy systems. Fermentative substrate is an important factor influencing the cost of butanol production. Lignocelluloses and hemicelluloses are the most abundant renewable resource on the planet, and have attracted much attention recently due to their potential as substrates for biofuel production. However, during the pretreatment of lignocelluloses to get fermentative substrates, a complex mixture of microbial inhibitors are generated as by-products. In this study, we investigated the effect of various inhibitory compounds, ferulic acid, syringaldehyde, ρ-coumaric acid, 4-hydroxybenzoic acid, vanilin acid, and vanilin, on butanol production and growth with Clostridium beijerinckii NCIMB 8052 and Clostridium acetobutylicum ATCC 824. The effect of the inhibitory compounds on the growth and production of butanol were monitored over 72-hour of batch cultivation with C. beijerinckii NCIMB 8052 and C. acetobutylicum ATCC 824. The exposure to ρ-coumaric acid led a reduced butanol production for both C. beijerinckii NCIMB 8052 and C. acetobutylicum ATCC 824, while the addition of syringaldehyde did not show any reduction on butanol production for both strains.

Covalent attachment, aggregate coat ing or entrapment of enzymes with nanof ibers yields highly act ive and stable systems for the applicat ions in the various f ields such ac bioconversion, biomedicals, and biosensors. In this study, we compared the stabilit ies of enzymes, which were immobilized by several nano-based biosilif icat ion. Glutathione Stransferase (GST) as the model enzyme are used, and GST and R5 tagged GST gene was amplif ied by PCR and inserted into the pET-28 a(+) vector (Novagen). It was expressed in E.coli BL21, and then purif ied with aff inity chromatography. GST was immobilized into the nanof iber consisted of polystyrene (PS) and poly (styrene-co-maleic anhydride) (PSMA). R5 catalyzed the format ion of silica nanopart icles (about 200 nm in diameter) with GST retained its act ivity.

The generation of acetone, butanol and ethanol (ABE) by C. acetobutylicum was monitored using batch cultures exposed to different concentrations of ferulic acid. It was found that 0.5 g/L ferulic acid led to an improved solventogenesis, with as much as a 6-fold increase in the production of butanol and acetone. It was also found that these cultures had a higher optical density, better glucose utilization and higher pH. Further tests were performed using structurally similar compounds, including coumaric acid and cinnamic acid, to better understand the mechanism leading to this enhances solventogenic activity. Initial results hint that hydroxymethyl group(s) present on the benzyl ring may play a significant role in this induction.

Biodiesel, which is a renewable, biodegradable, nontoxic fuel and similar to diesel fuels, can be used directly for diesel engines. Biodiesel is usually produced by transesterification of vegetable oils and animal fats with alcohols over various catalysts and has received considerable attention for several years. Canola seed is one of the major vegetable oil resources for biodiesel production due to its high oil content. In this work, we investigated a transesterification of canola oil with methanol in the presence of a immobilized lipase (Novozyme 435). The reaction was conducted under ultrasound-assisted supercritical conditions using carbon dioxide as solvent in a batch reactor. An enzymatic transesterification method of canola oil in ultrasound-assisted supercritical carbon dioxide (SCCO2) has been developed that allows a short reaction time and high yield through enhanced mass transfer in SCCO2 by ultrasonication. The effects of various parameters such as enzyme loading, methanol to oil molar ratio, reaction time, and ultrasonication power were investigated in this reaction system.

Enzyme activity is an important character because industry requires efficient catalysts. In this study, bacteriophage T4 lysozyme was selected as a model enzyme and the flexibility of the mutants was analyzed to understand the mechanism of activity enhancement. The group having enhanced activity showed a tendency that the changed residues in the mutants were, in many cases, Glu, Asp and many of them were located in the helix edge. Because the flexibility is related with the enzyme motion and this can affect the catalytic activity of the enzyme, spring model for enzyme deformation was proposed for the calculation of residual force. And torques at the edges were calculated also. This new modeling method found that the most twisted region of T4 lysozyme was R80, already known as the hinge-bending region. Thus, this model equation has the validity to guess the enzyme motion. And it was found that the mutations at the edges located far away from R80 increases enzyme activity. This is the clue that activity enhancement could be achieved by the introduction of hydrophilic residues at some appropriate helix edges in case of hydrolases doing hinge-bending motion during catalysis.

Lipases are used as efficient biocatalysts in many industrial processes. Especially, Candida antarctica lipase B (Cal B) draws attention because it can be used for stereoselective transformations and polymer synthesis, and the potential process for biodiesel production using lipase instead of chemical catalysts is under development. Cal B was expressed in the methylotrophic yeast P ichia pastoris X-33 using pPICZαB vector, which has α-factor, helping protein secretion by methanol induction. The colonies showing a halo were selected, and the expression and secretion of Cal B in flask cultivation with glycerol as a carbon source and methanol induction was confirmed by SDS-PAGE and the activity test using p-NPP. A 5-l fermentor was used to investigate the effects of cultivation conditions on the lipase expression and secretion.

3-Hydroxypropionaldehyde (3-HPA), an intermediary compound of glycerol metabolism in bacteria, serves as a precursor to 3-Hydroxypropionic acid (3-HP), a commercially valuable platform chemical. To achieve the effective conversion of 3-HPA to 3-HP, an aldH gene encoding an aldehyde dehydrogenase in Escherichia coli K-12 (AldH) was cloned, expressed, and characterized for its properties. The recombinant AldH exhibited broad substrate specificity for various aliphatic and aromatic aldehydes. AldH preferred NAD+ over NADP+ as a cofactor for the oxidation of most aliphatic aldehydes tested. Among the aldehydes used, the specific activity was highest (38.1 U mg−1 protein) for 3-HPA at pH 8.0 and 37 °C. The catalytic efficiency (kcat) and the specificity constant (kcat/Km) for 3-HPA in the presence of NAD+ were 28.5 s−1 and 58.6×103 M−1 s−1, respectively. The AldH activity was enhanced in the presence of disulfide reductants such as dithiothreitol (DTT) or 2-mercaptoethanol, while several metal ions, particularly Hg2+, Ag+, Cu2+, and Zn2+, inhibited AldH activity. This study illustrates that AldH is a potentially useful enzyme in converting 3-HPA to 3-HP.

Ketoprofen,anon-steroidalanti-inflammatorydrug, inhibits the synthesis of prostaglandin. Previously, a novel hydrolase(Est25) with high ketoprofen specificity was identified using a metagenomic library from environmental samples. Interestingly, the primary sequence of Est25 shows similarities to members of the hormone-sensitive lipase (HSL) family such as brefeldin A esterase (BFAE, 48% identity) from B acillus subtilis, a carboxylesterase (AFEST, 33%) from Archaeoglobus fulgidus, and a carboxyesterase (EST2, 30%) from Alicyclobacillus acidocaldarius. The HSL family is a class of enzymes that mobilize fatty acids for energy metabolism in adipose tissue. Although the members of the HSL family share an α/β-hydrolase fold, each structure has its own arrangement of α-helices and β-sheets. Therefore, the crystal structure of Est25 is essential to understand the enzyme’s catalytic mechanism and substrate specificities for (R,S)-ketoprofen at the molecular level. In addition, the Est25 crystal structure is expected to provide structural and functional insights into the HSL family. Est25 was crystallized from 2.4M sodium malonate, pH7.0, and X-ray diffraction data to 1.49 Å were collected using synchrotron radiation. The crystals belong to the monoclinic space group C2 with unit cell parameters a = 197.8 Å , b = 95.2 Å , c = 99.4 Å , and β = 97.1˚.

The thiol-disulfide oxidoreductase thioredoxin-1 (TRX) is known to be secreted by leukocytes and to exhibit cytokine-like properties. Extracellular effects of TRX require a functional active site, suggesting a redox-based mechanism of action. However, specific cell surface proteins and pathways coupling extracellular TRX redox activity to cellular responses have not been identified so far. We first found that exogenous TRX not only scavenger but also effected intracellular proteins using proteomic approaches. Therefore, the effect of a thiol-antioxidant protein TRX have appropriated on regulation in a human skin malignant melanoma cell line (A375). Mechanism-based Histidine pull down and immunoprecipitation technique used to identify TRX binding partner proteins. We found that up/down regulated intracellular proteins were identified under the condition of LPS-treated A375 cells and co-treat with exogenous TRX using in vitro 2-DE proteomic approaches. As well as, confirmed in vivo mouse inflammation models used to quantitatively measure IL-6, IL-10, MCP-1 and TNF protein levels in serum samples. Our studies demonstrate that exogenous TRX has anti-inflammatory properties and intracellular regulator activity in vivo and in vitro. These results have a therapeutic role in skin inflammation therapy.

In this work is reported the urease immobilized on magnetically separable polyaniline nanofibers to improve the enzyme activity. Immobilized urease shows good activity as 5times more than free urease. The immobilized urease showed improved stability and recyclability. PAMP-urease showed only a 3~5% decrease in activity after 1 months of vigorous shaking conditions (200 rpm) and at room temperature (25oC). After 10 times recycle, the residual activities of PAMP-urease was 92%.

Polyanilie based nanomaterials and which were magnetically separable have been used for the immobilization of enzymes for recovery and recycle. Polyaniline nanomaterials which mixed with iron oxide nanoparticle were shown distinctive differences in morphology and revealed aggregated shapes. BET surface area was 46.36m2 g-1 decreased with the nanoparticle addition from 58.37m2 g-1. The magnetically separable catalase enzymes were developed using cross linked enzyme aggregation on the surface of nanomaterials. Developed nanoenzymes were used for the bioconversion of hydrogen peroxide and were successfully recovered with magnet and used repeatedly for reaction. The residual activity after 5 times reuse was 88% of its original activity.

We have identified twenty-first aminoacyl-tRNA synthetase-suppressor tRNA pairs for possible use in site-specific incorporation of amino acid analogues into proteins in eukaryotes (E. coli glutaminyl-tRNA synthetase (GlnRS) / suppressor tRNAs derived from E. coli tRNAGln or human initiator tRNA). Our goal is to identify mutants of E. coli GlnRS that can utilize the keto-containing glutamine analogue 2-amino-5-ketohexanoic acid (wherein the amide side chain of glutamine is replaced by a methyl ketone). The keto group can undergo a variety of reactions with hydrazides, alkoxyamines, and semicarbazides under physiological conditions. It can be used to produce proteins modified with fluorescent group, biotin, polyethylenglycol, saccharide and so on. To genetically encode 2-amino-5-ketohexanoic acid in yeast, We are working on isolation of the desired mutants of E. coli GlnRS using rational design and random mutagenesis of the active site.

D-Psicose, a rare keto-hexose, are not abundant in nature and are difficult to prepare by chemical methods. D-Psicose has sweet taste like general sugars, but calorie is near to zero because it is not metabolized in the human body. For the production of D-psicose, we used allulose-6-phosphate 3-epimerase of Escherichia coli and D-tagatose 3-epimerase of Agrobacterium tumefeciens. In order to use the allulose-6-phosphate 3-epimerase, E. coli was transformed with pTalsE containing alsE. Fructose-6-phosphate of glycolysis pathway might be epimerized to psicose-6-phosphate by alsE. The pfkA gene encoding 6-phosphofructokinase of glycolysis was deleted to make the psicose producing E. coli blocking use of fructose, and allose operon, including alsE, alsI, alsK, alsA, alsB, alsC and alsR genes, was deleted. When E. coli (
pfkA, als2) harboring pTalsE was cultivated in 2YT medium with 30g/ℓ of glucose and fructose, 0.5 and 1g/ℓ of D-psicose was obtained approximately, respectively. The other scheme for production of D-psicose was performed with D-tagatose 3-epimerase by which fructose was epimerized to D-psicose. When recombinant E. coli (
pfkA, als2) harboring pTPE with Atu4750 was cultivated in 2YT medium containing 30g/ℓ of fructose, D-psicose was obtained upto 6g/ℓ. This work was supported by the 21C Frontier Microbial Genomics and Applications Center Program, EB-NCRC (Grant No. R15-2003-012-02001-0), Brain Pool program and BK21 program of Korea.

CiP(Coprinus cinereus peroxidase) catalyzing a phenolic derivates such as m-cresol, p-cresol, bisphenol A can produce a potential applicable materials in electrooptics. For application to the industry, to know the properties of polymer and its behavior control is significantly import. We investigate in the polymerization of m-cresol and p-cresol using CiP as catalyst effects of organic solvent composition, yield, solubility and molecular weight. It has a difference with solubility, ratio of phenylene bridge, yield to perform m-cresol and p-cresol synthesized in the same conditions(pH, solvent composition, temperature). Unexpectedly, increasing of p-cresol ratio causes the decreasing of yield, molecular weight and ratio of oxyphenylen bridge. By the contrast, increasing of m-cresol causes the increasing of yield, molecular weight and ratio of oxyphenylen bridge, however it has a increasing insoluble product. To improve their a individual defect is overcome by copolymerization of m-cresol and p-cresol ,using their complementary property, from the appropriable water miscible organic solvent and buffer.

The electrochemical regeneration of nicotinamide cofactor is of particular interest in oxidoreductase catalyzed reactions such as chemical synthesis, degradation of pollutant, construction of biosensor, and fuel generation. Because most of oxidoreductase enzymes require expensive cofactors for catalysis, the cofactors have to be regenerated. For the efficient cofactor regeneration under low overpotential, indirect methods utilizing mediators have been frequently used. Reactions using mediators can occurred under low overpotential and prevent electrode fouling, but some of these mediators are toxic to the enzymes. Also, the stability of the mediator can affect the whole reaction performance. Therefore, construction of mediator-free cofactor regeneration system is very attractive and necessary. In this study, we screened several kinds of new metal oxides as an electrode material for direct electrochemical cofactor regeneration. Using these new metal oxide electrode, cofactors were regenerated efficiently under low overpotential without using mediator. Details will be presented and discussed.