Kim HK

References (40)

Title : Synthesis of Isoamyl Fatty Acid Ester, a Flavor Compound, by Immobilized Rhodococcus Cutinase - Jeon_2024_J.Microbiol.Biotechnol_34_1
Author(s) : Jeon YW , Song HM , Lee KY , Kim YA , Kim HK
Ref : J Microbiol Biotechnol , 34 :1 , 2024
Abstract : Isoamyl fatty acid esters (IAFEs) are widely used as fruity flavor compounds in the food industry. In this study, various IAFEs were synthesized from isoamyl alcohol and various fatty acids using a cutinase enzyme (Rcut) derived from Rhodococcus bacteria. Rcut was immobilized on methacrylate divinylbenzene beads and used to synthesize isoamyl acetate, butyrate, hexanoate, octanoate, and decanoate. Among them, Rcut synthesized isoamyl butyrate (IAB) most efficiently. Docking model studies showed that butyric acid was the most suitable substrate in terms of binding energy and distance from the active site serine (Ser114) gamma-oxygen. Up to 250 mM of IAB was synthesized by adjusting reaction conditions such as substrate concentration, reaction temperature, and reaction time. When the enzyme reaction was performed by reusing the immobilized enzyme, the enzyme activity was maintained at least six times. These results demonstrate that the immobilized Rcut enzyme can be used in the food industry to synthesize a variety of fruity flavor compounds, including IAB.
ESTHER : Jeon_2024_J.Microbiol.Biotechnol_34_1
PubMedSearch : Jeon_2024_J.Microbiol.Biotechnol_34_1
PubMedID: 38754998
Gene_locus related to this paper: rhofa-a0a7i8e2z4

Title : Synthesis of New Multifunctional Linolenic Acid Vanillyl Ester and Investigation of Antioxidant and Antibacterial Activities - Marvella_2024_Appl.Biochem.Biotechnol__
Author(s) : Marvella JI , Kim HK
Ref : Appl Biochem Biotechnol , : , 2024
Abstract : Vanillyl alcohol (VA) possesses potent antioxidant activity, yet its applicability is hindered by its limited solubility in emulsions or non-polar organic solvents. Conversely, long-chain polyunsaturated fatty acids exhibit antibacterial properties. The combination of these compounds offers the prospect of developing novel phenolic lipid compounds with dual antioxidant and antibacterial activities, alongside enhanced solubility capabilities. In this investigation, linolenic acid vanillyl ester (LAVE) was synthesized from VA and linseed oil (LO) through a transesterification reaction employing immobilized lipase. Optimization of LAVE production was achieved by varying reaction temperature, substrate concentration, and reaction time. LAVE demonstrated efficacy in scavenging both 2,2-diphenyl-1-picryhydrazyl and 2,2'-azino-bis (3-ethylbenthiazoline-6-sulphonic acid) radicals in organic solvents. Antioxidant testing via lipid oxidation analysis revealed that LAVE, when distributed within emulsions, effectively impeded the formation of conjugated dienes and conjugated trienes. Furthermore, LAVE exhibited antibacterial activity against four strains of spoilage bacteria: Bacillus subtilis, Bacillus coagulans, Pseudomonas fluorescens, and Alcaligenes faecalis. Zeta potential analysis substantiated the binding of LAVE to the bacterial cell surface. Propidium iodide uptake assay and fluorescence microscopy further elucidated that LAVE induces cell lysis by augmenting membrane permeability.
ESTHER : Marvella_2024_Appl.Biochem.Biotechnol__
PubMedSearch : Marvella_2024_Appl.Biochem.Biotechnol__
PubMedID: 38647998

Title : Cloning and characterization of an acidic lipase from a lipolytic bacterium in tempeh - Nur_2023_J.Genet.Eng.Biotechnol_21_157
Author(s) : Nur N , Suwanto A , Meryandini A , Suhartono MT , Puspitasari E , Kim HK
Ref : J Genet Eng Biotechnol , 21 :157 , 2023
Abstract : BACKGROUND: Lipases have emerged as essential biocatalysts, having the ability to contribute to a wide range of industrial applications. Microbial lipases have garnered significant industrial attention due to their stability, selectivity, and broad substrate specificity. In the previous study, a unique lipolytic bacterium (Micrococcus luteus EMP48-D) was isolated from tempeh. It turns out the bacteria produce an acidic lipase, which is important in biodiesel production. Our main objectives were to clone the acidic lipase and investigate its potential in biodiesel production. RESULT: In this study, the gene encoding a lipase from M. luteus EMP48-D was cloned and expressed heterologously in Escherichia coli. To our knowledge, this is the first attempt at the cloning and expression of the lipase gene from Micrococcus luteus. The amino acid sequence was deduced from the nucleotide sequence (1356 bp) corresponded to a protein of 451 amino acid residues with a molecular weight of about 40 kDa. The presence of a signal peptide suggested that the protein was extracellular. A sequence analysis revealed that the protein had a lipase-specific Gly-X-Ser-X-Gly motif. The enzyme was identified as an acidic lipase with a pH preference of 5.0. Fatty acid preferences for enzyme activities were C8 and C12 (p-nitrophenyl esters), with optimum temperatures at 30-40 degreesC and still remaining active at 80 degreesC. The enzyme was also shown to convert up to 70% of the substrate into fatty acid methyl ester. CONCLUSION: The enzyme was a novel acidic lipase that demonstrated both hydrolytic and transesterification reactions. It appeared particularly promising for the synthesis of biodiesel as this enzyme's catalytic reaction was optimum at low temperatures and was still active at high temperatures.
ESTHER : Nur_2023_J.Genet.Eng.Biotechnol_21_157
PubMedSearch : Nur_2023_J.Genet.Eng.Biotechnol_21_157
PubMedID: 38038870
Gene_locus related to this paper: miclu-a0a5b8ywx7

Title : Evogliptin, a DPP-4 inhibitor, prevents diabetic cardiomyopathy by alleviating cardiac lipotoxicity in db\/db mice - Pham_2023_Exp.Mol.Med__
Author(s) : Pham TK , Nguyen THT , Yi JM , Kim GS , Yun HR , Kim HK , Won JC
Ref : Exp Mol Med , : , 2023
Abstract : Dipeptidyl peptidase-4 (DPP-4) inhibitors are glucose-lowering drugs for type 2 diabetes mellitus (T2DM). We investigated whether evogliptin(a) (EVO), a DPP-4 inhibitor, could protect against diabetic cardiomyopathy (DCM) and the underlying mechanisms. Eight-week-old diabetic and obese db/db mice were administered EVO (100 mg/kg/day) daily by oral gavage for 12 weeks. db/db control mice and C57BLKS/J as wild-type (WT) mice received equal amounts of the vehicle. In addition to the hypoglycemic effect, we examined the improvement in cardiac contraction/relaxation ability, cardiac fibrosis, and myocardial hypertrophy by EVO treatment. To identify the mechanisms underlying the improvement in diabetic cardiomyopathy by EVO treatment, its effect on lipotoxicity and the mitochondrial damage caused by lipid droplet accumulation in the myocardium were analyzed. EVO lowered the blood glucose and HbA1c levels and improved insulin sensitivity but did not affect the body weight or blood lipid profile. Cardiac systolic/diastolic function, hypertrophy, and fibrosis were improved in the EVO-treated group. EVO prevented cardiac lipotoxicity by reducing the accumulation of lipid droplets in the myocardium through suppression of CD36, ACSL1, FABP3, PPARgamma, and DGAT1 and enhancement of the phosphorylation of FOXO1, indicating its inhibition. The EVO-mediated improvement in mitochondrial function and reduction in damage were achieved through activation of PGC1a/NRF1/TFAM, which activates mitochondrial biogenesis. RNA-seq results for the whole heart confirmed that EVO treatment mainly affected the differentially expressed genes (DEGs) related to lipid metabolism. Collectively, these findings demonstrate that EVO improves cardiac function by reducing lipotoxicity and mitochondrial injury and provides a potential therapeutic option for DCM.
ESTHER : Pham_2023_Exp.Mol.Med__
PubMedSearch : Pham_2023_Exp.Mol.Med__
PubMedID: 37009790

Title : Production of octyl butyrate using psychrophilic mutant lipase from Croceibacter atlanticus LipCA lipase developed by a molecular evolution technique - Song_2023_Enzyme.Microb.Technol_173_110370
Author(s) : Song HM , Kim HK
Ref : Enzyme Microb Technol , 173 :110370 , 2023
Abstract : Lipases are used to synthesize a variety of industrially useful compounds. Among them, psychrophilic lipase can be used to synthesize thermo-labile compounds at low temperatures. In this study, random mutagenesis was introduced into Antarctic Croceibacter atlanticus lipase gene using error-prone PCR, resulting in changes in its protein sequence. Through two rounds of mutagenesis and screening, we found that a mutant R1 showed an enhanced activity at low temperatures. Mutant R1 had five mutations (F43L, S48G, S49G, D141K, and K297R) and higher k(cat)/K(M) value than the wild type (WT) at 10 degreesC. We immobilized this enzyme on methacrylate divinylbenzene resin and used it to synthesize octyl butyrate, a flavor compound. The esterification reaction proceeded even at 10 degreesC. Mutant R1 synthesized the ester compound faster than the WT. To determine which amino acids were responsible for the increase of activity, site-directed mutagenesis was performed to introduce five back mutations into mutant R1. Three back mutants (L43F, G48S, G49S) showed significant decreases of activity at low temperatures, indicating that these amino acids were closely related to the increase in activity. This psychrophilic mutant R1 is expected to be used in low-temperature enzyme conversion reactions in the food industry.
ESTHER : Song_2023_Enzyme.Microb.Technol_173_110370
PubMedSearch : Song_2023_Enzyme.Microb.Technol_173_110370
PubMedID: 38043250
Gene_locus related to this paper: 9gamm-b4x2p5

Title : Functional production, characterization, and immobilization of a cold-adapted cutinase from Antarctic Rhodococcus sp - Won_2022_Protein.Expr.Purif__106077
Author(s) : Won SJ , Yim JH , Kim HK
Ref : Protein Expr Purif , :106077 , 2022
Abstract : A lipolytic enzyme (Rcut) was discovered from the Rhodococcusstrain (RosL12) isolated from the Antarctic Ross Sea. The corresponding gene composed of 651 bases encoding 216 amino acids. It was found to be a cutinase gene through BLAST search. Rcut has a signal sequence consisting of 29 amino acids. An active Rcut was produced after the intact gene containing the signal sequence was transformed into Escherichia coli Rosetta-gami 2 (DE3) pLysS. Rcut was purified through a nickel-nitrilotriacetic acid purification system and a carboxymethyl Sepharose column chromatography. Its specific activity was 2190 U/mg. Rcut showed the highest activity at 40 degreesC and had a low activation energy of 3.16 kcal/mol. This means that it is a typical cold-adapted enzyme. Rcut showed high activity towards medium chain fatty acids (C(4)-C(10)). Rcut degraded polycaprolactone and polyethylene terephthalate, suggesting that it could be used for decomposition of synthetic plastics causing environmental pollution. Rcut was immobilized on methacrylate-divinyl benzene bead. This immobilized Rcut (immRcut) showed higher thermal stability than the free enzyme. ImmRcut performed transesterification of various esters and ethanol in a non-polar solvent, suggesting that it could be used for the synthesis of industrially useful ester compounds.
ESTHER : Won_2022_Protein.Expr.Purif__106077
PubMedSearch : Won_2022_Protein.Expr.Purif__106077
PubMedID: 35314296
Gene_locus related to this paper: rhofa-a0a7i8e2z4

Title : Synthesis of Short-Chain Alkyl Butyrate through Esterification Reaction Using Immobilized Rhodococcus Cutinase and Analysis of Substrate Specificity through Molecular Docking - Won_2022_J.Microbiol.Biotechnol_33_1
Author(s) : Won SJ , Yim JH , Kim HK
Ref : J Microbiol Biotechnol , 33 :1 , 2022
Abstract : Alkyl butyrate with fruity flavor is known as an important additive in the food industry. We synthesized various alkyl butyrates from various fatty alcohol and butyric acid using immobilized Rhodococcus cutinase (Rcut). Esterification reaction was performed in a non-aqueous system including heptane, isooctane, hexane, and cyclohexane. As a result of performing the alkyl butyrate synthesis reaction using alcohols of various chain lengths, it was found that the preference for the alcohol substrate had the following order: C6 > C4 > C8 > C10 > C2. Through molecular docking analysis, it was found that the greater the hydrophobicity of alcohol, the higher the accessibility to the active site of the enzyme. However, since the number of torsions increased as the chain length increased, it became difficult for the hydroxyl oxygen of the alcohol to access the gamma O of serine at the enzyme active site. These molecular docking results were consistent with substrate preference results of the Rcut enzyme. The Rcut maintained the synthesis efficiency at least for 5 days in isooctane solvent. We synthesized as much as 452 mM butyl butyrate by adding 100 mM substrate daily for 5 days and performing the reaction. These results show that Rcut is an efficient enzyme for producing alkyl butyrate used in the food industry.
ESTHER : Won_2022_J.Microbiol.Biotechnol_33_1
PubMedSearch : Won_2022_J.Microbiol.Biotechnol_33_1
PubMedID: 36524336
Gene_locus related to this paper: rhofa-a0a7i8e2z4

Title : Antioxidant and Antibacterial Activity of Caprylic Acid Vanillyl Ester Produced by Lipase-Mediated Transesterification - Kim_2021_J.Microbiol.Biotechnol_31_317
Author(s) : Kim JJ , Kim HK
Ref : J Microbiol Biotechnol , 31 :317 , 2021
Abstract : Vanillyl alcohol (VA), which is abundant in Vanilla bean, has strong antioxidant activity. However, the use of VA in the food and cosmetics industries is limited, due to its low solubility in emulsion or organic solvents. Meanwhile, medium chain fatty acids and medium chain monoglycerides have antibacterial activity. We synthesized butyric acid vanillyl ester (BAVE) or caprylic acid vanillyl ester (CAVE) from VA with tributyrin or tricaprylin through transesterification reaction using immobilized lipases. BAVE and CAVE scavenged 2,2-diphenyl-1-picrylhydrazyl radicals in organic solvents. In addition, BAVE and CAVE decreased the production rate of conjugated diene and triene in the menhaden oil-in-water emulsion system. While BAVE showed no antibacterial activity, CAVE showed antibacterial activity against food spoilage bacteria, including Bacillus coagulans. In this study, the antibacterial activity of vanillyl ester with medium chain fatty acid was first revealed. Zeta potential measurements confirmed that BAVE and CAVE were inserted into B. coagulans membrane. In addition, the propidium iodide uptake assay and fluorescent microscopy showed that CAVE increased B. coagulans membrane permeability. Therefore, CAVE is expected to play an important role in the food and cosmetics industries as a bi-functional material with both antioxidant and antibacterial activities.
ESTHER : Kim_2021_J.Microbiol.Biotechnol_31_317
PubMedSearch : Kim_2021_J.Microbiol.Biotechnol_31_317
PubMedID: 33203820

Title : Increased mRNA stability and expression level of Croceibacter atlanticus lipase gene developed through molecular evolution process - Jeong_2021_J.Microbiol.Biotechnol_31_
Author(s) : Jeong HB , Kim HK
Ref : J Microbiol Biotechnol , 31 : , 2021
Abstract : In order to use an enzyme industrially, it is necessary to increase the activity of the enzyme and optimize the reaction characteristics through molecular evolution techniques. We used the error-prone PCR method to improve the reaction characteristics of LipCA lipase discovered in Antarctic Croceibacter atlanticus. Recombinant Escherichia coli colonies showing large halo zones were selected in tributyrin-containing medium. The lipase activity of one mutant strain (M3-1) was significantly increased, compared to the wild-type (WT) strain. M3-1 strain produced about three times more lipase enzyme than did WT strain. After confirming the nucleotide sequence of the M3-1 gene to be different from that of the WT gene by four bases (73, 381, 756, and 822), the secondary structures of WT and M3-1 mRNA were predicted and compared by RNAfold web program. Compared to the mean free energy (MFE) of WT mRNA, that of M3-1 mRNA was lowered by 4.4 kcal/mol, and the MFE value was significantly lowered by mutations of bases 73 and 756. Site-directed mutagenesis was performed to find out which of the four base mutations actually affected the enzyme expression level. Among them, one mutant enzyme production decreased as WT enzyme production when the base 73 was changed (T->C). These results show that one base change at position 73 can significantly affect protein expression level, and demonstrate that changing the mRNA sequence can increase the stability of mRNA, and can increase the production of foreign protein in E. coli.
ESTHER : Jeong_2021_J.Microbiol.Biotechnol_31_
PubMedSearch : Jeong_2021_J.Microbiol.Biotechnol_31_
PubMedID: 34024893

Title : Standardized Extract (HemoHIM) Protects against Scopolamine-Induced Amnesia in a Murine Model - Kim_2021_Evid.Based.Complement.Alternat.Med_2021_8884243
Author(s) : Kim SK , Kwon DA , Kim YS , Lee HS , Kim HK , Kim WK
Ref : Evid Based Complement Alternat Med , 2021 :8884243 , 2021
Abstract : HemoHIM is a medicinal herbal preparation of Angelica gigas Nakai (Apiaceae), Cnidium officinale Makino (Umbelliferae), and Paeonia lactiflora Pallas (Paeoniaceae) designed for immune regulation. In the present study, the memory-enhancing effects of a standardized extract (HemoHIM) on scopolamine-induced memory impairment in a murine model was investigated. To induce amnesia, scopolamine (1 mg/kg) was intraperitoneally (i.p.) injected into mice 30 min before the start of behavioral tests. The Y-maze, novel object recognition test (NORT), and passive avoidance task (PAT) were used to evoke memory functions. HemoHIM significantly improved scopolamine-induced memory impairment in ICR mice, which was evidenced by an improvement of spontaneous alternation in the Y-maze, recognition index in NORT, and latency time in PAT. To elucidate the possible mechanism, the cholinergic activity and mRNA levels of choline acetyltransferase (ChAT), muscarinic acetylcholine receptor (mAchR), brain-derived neurotrophic factor (BDNF), and cAMP response element-binding protein (CREB) were measured using reverse transcription (RT-PCR) and western blot analyses, respectively. HemoHIM treatment attenuated the scopolamine-induced hyperactivation of acetylcholinesterase (AchE) activity. In addition, ChAT, mAchR, and CREB mRNA levels were increased in the hippocampus compared with the scopolamine group. Furthermore, HemoHIM treatment resulted in elevated BDNF protein expression. These results indicate that HemoHIM may exert antiamnesic activity by increasing Ach and inhibiting AchE in the hippocampus. In addition, HemoHIM has therapeutic potential by upregulating ChAT, mAchR, and BDNF, which is apparently mediated by activation of the CREB and ERK signaling pathways.
ESTHER : Kim_2021_Evid.Based.Complement.Alternat.Med_2021_8884243
PubMedSearch : Kim_2021_Evid.Based.Complement.Alternat.Med_2021_8884243
PubMedID: 33815562

Title : Isolation and biochemical characterization of Bacillus pumilus lipases from the Antarctic - Arifin_2013_J.Microbiol.Biotechnol_23_661
Author(s) : Arifin AR , Kim SJ , Yim JH , Suwanto A , Kim HK
Ref : J Microbiol Biotechnol , 23 :661 , 2013
Abstract : Lipase-producing bacterial strains were isolated from Antarctic soil samples using the tricaprylin agar plate method. Seven strains with relatively strong lipase activities were selected. All of them turned out to be Bacillus pumilus strains by the 16S rRNA gene sequence analysis. Their corresponding lipase genes were cloned, sequenced, and compared. Finally, three different Bacillus pumilus lipases (BPL1, BPL2, and BPL3) were chosen. Their amino acid sequence identities were in the range of 92-98% with the previous Bacillus pumilus lipases. Their optimum temperatures and pHs were measured to be 40 degrees C and pH 9. Lipase BPL1 and lipase BPL2 were stable up to 30 degrees C, whereas lipase BPL3 was stable up to 20 degrees C. Lipase BPL2 was stable within a pH range of 6-10, whereas lipase BPL1 and lipase BPL3 were stable within a pH range of 5-11, showing strong alkaline tolerance. All these lipases exhibited high hydrolytic activity toward pnitrophenyl caprylate (C8). In addition, lipase BPL1 showed high hydrolytic activity toward tributyrin, whereas lipase BPL2 and lipase BPL3 hydrolyzed tricaprylin and castor oil preferentially. These results demonstrated that the three Antarctic Bacillus lipases were alkaliphilic and had a substrate preference toward short- and mediumchain triglycerides. These Antarctic Bacillus lipases might be used in detergent and food industries.
ESTHER : Arifin_2013_J.Microbiol.Biotechnol_23_661
PubMedSearch : Arifin_2013_J.Microbiol.Biotechnol_23_661
PubMedID: 23648856

Title : Expression and biochemical characterization of cold-adapted lipases from Antarctic Bacillus pumilus strains - Litantra_2013_J.Microbiol.Biotechnol_23_1221
Author(s) : Litantra R , Lobionda S , Yim JH , Kim HK
Ref : J Microbiol Biotechnol , 23 :1221 , 2013
Abstract : Two lipase genes (bpl1 and bpl3) from Antarctic Bacillus pumilus strains were expressed in Bacillus subtilis. Both recombinant lipases BPL1 and BPL2 were secreted to the culture medium and their activities reached 3.5 U/ml and 5.0 U/ml, respectively. Their molecular masses apparent using SDS-PAGE were 23 kDa for BPL1 and 19 kDa for BPL3. Both lipases were purified to homogeneity using ammonium sulfate precipitation and HiTrap SP FF column and Superose 12 column chromatographies. The final specific activities were estimated to be 328 U/mg for BPL1 and 310 U/mg for BPL3. Both lipases displayed an optimum temperature of 35 degreesC, similar to other mesophilic enzymes. However, they maintained as much as 70% and 80% of the maximum activities at 10 degreesC. Accordingly, their calculated activation energy at a temperature range of 10-35 degreesC was 5.32 kcal/mol for BPL1 and 4.26 kcal/mol for BPL3, typical of cold-adapted enzymes. The optimum pH of BPL1 and BPL3 was 8.5 and 8.0, respectively, and they were quite stable at pH 7.0-11.0, showing their strong alkaline tolerance. Both lipases had a preference toward medium chain length (C6-C10) fatty acid substrates. These results indicate the potential for the two Antarctic B. pumilus lipases as catalysts in bioorganic synthesis, food, and detergent industries.
ESTHER : Litantra_2013_J.Microbiol.Biotechnol_23_1221
PubMedSearch : Litantra_2013_J.Microbiol.Biotechnol_23_1221
PubMedID: 23770563

Title : Gene cloning and characterization of a cold-adapted esterase from Acinetobacter venetianus V28 - Kim_2012_J.Microbiol.Biotechnol_22_1245
Author(s) : Kim YO , Heo YL , Kim HK , Nam BH , Kong HJ , Kim DG , Kim WJ , Kim BS , Jee YJ , Lee SJ
Ref : J Microbiol Biotechnol , 22 :1245 , 2012
Abstract : Acinetobacter venetians V28 was isolated from the intestine of righteye flounder, Poecilopsetta plinthus caught in Vietnam seawater, and the esterase gene was cloned using a shotgun method. The amino acid sequence deduced from the nucleotide sequence (1,017 bp) corresponded to a protein of 338 amino acid residues with a molecular weight of 37,186. The esterase had 87% and 72% identities with the lipases of A. junii SH205 and A. calcoaceticus RUH2202, respectively. The esterase contained a putative leader sequence, as well as the conserved catalytic triad (Ser, His, Asp), consensus pentapeptide GXSXG, and oxyanion hole sequence (HG). The protein from the strain V28 was produced in both a soluble and an insoluble form when the Escherichia coli cells harboring the gene were cultured at 18 degrees C. The maximal activity of the purified enzyme was observed at a temperature of 40 degrees C and pH 9.0 using p-NP-caprylate as substrate; however, relative activity still reached to 70% even at 5 degrees C with an activation energy of 3.36 kcal/mol, which indicated that it was a cold-adapted enzyme. The enzyme was a nonmetalloprotein and was active against p-nitrophenyl esters of C4, C8, and C14. Remarkably, this enzyme retained much of its activity in the presence of commercial detergents and organic solvents. This cold-adapted esterase will be applicable as catalysts for reaction in the presence of organic solvents and detergents.
ESTHER : Kim_2012_J.Microbiol.Biotechnol_22_1245
PubMedSearch : Kim_2012_J.Microbiol.Biotechnol_22_1245
PubMedID: 22814499

Title : Gene cloning and catalytic characterization of cold-adapted lipase of Photobacterium sp. MA1-3 isolated from blood clam - Kim_2012_J.Biosci.Bioeng_114_589
Author(s) : Kim YO , Khosasih V , Nam BH , Lee SJ , Suwanto A , Kim HK
Ref : J Biosci Bioeng , 114 :589 , 2012
Abstract : A lipase-producing Photobacterium strain (MA1-3) was isolated from the intestine of a blood clam caught at Namhae, Korea. The lipase gene was cloned by shotgun cloning and encoded 340 amino acids with a molecular mass of 38,015 Da. It had a very low sequence identity with other bacterial lipases, with the exception of that of Photobacterium lipolyticum M37 (83.2%). The MA1-3 lipase was produced in soluble form when Escherichia coli cells harboring the gene were cultured at 18 degrees C. Its optimum temperature and pH were 45 degrees C and pH 8.5, respectively. Its activation energy was calculated to be 2.69 kcal/mol, suggesting it to be a cold-adapted lipase. Its optimum temperature, temperature stability, and substrate specificity were quite different from those of M37 lipase, despite the considerable sequence similarities. Meanwhile, MA1-3 lipase performed a transesterification reaction using olive oil and various alcohols including methanol, ethanol, 1-propanol, and 1-butanol. In the presence of t-butanol as a co-solvent, this lipase produced biodiesel using methanol and plant or waste oils. The highest biodiesel conversion yield (73%) was achieved using waste soybean oil and methanol at a molar ratio of 1:5 after 12 h using 5 units of lipase.
ESTHER : Kim_2012_J.Biosci.Bioeng_114_589
PubMedSearch : Kim_2012_J.Biosci.Bioeng_114_589
PubMedID: 22841866

Title : Characterization of lipases from Staphylococcus aureus and Staphylococcus epidermidis isolated from human facial sebaceous skin - Xie_2012_J.Microbiol.Biotechnol_22_84
Author(s) : Xie W , Khosasih V , Suwanto A , Kim HK
Ref : J Microbiol Biotechnol , 22 :84 , 2012
Abstract : Two staphylococcal lipases were obtained from Staphylococcus epidermidis S2 and Staphylococcus aureus S11 isolated from sebaceous areas on the skin of the human face. The molecular mass of both enzymes was estimated to be 45 kDa by SDS-PAGE. S2 lipase displayed its highest activity in the hydrolysis of olive oil at 32 degrees C and pH 8, whereas S11 lipase showed optimal activity at 31 degrees C and pH 8.5. The S2 lipase showed the property of cold-adaptation, with activation energy of 6.52 kcal/mol. In contrast, S11 lipase's activation energy, at 21 kcal/mol, was more characteristic of mesophilic lipases. S2 lipase was stable up to 45 degrees C and within the pH range from 5 to 9, whereas S11 lipase was stable up to 50 degrees C and from pH 6 to 10. Both enzymes had high activity against tributyrin, waste soybean oil, and fish oil. Sequence analysis of the S2 lipase gene showed an open reading frame of 2,067 bp encoding a signal peptide (35 aa), a pro-peptide (267 aa), and a mature enzyme (386 aa); the S11 lipase gene, at 2,076 bp, also encoded a signal peptide (37 aa), pro-peptide (255 aa), and mature enzyme (399 aa). The two enzymes maintained amino acid sequence identity of 98-99% with other similar staphylococcal lipases. Their microbial origins and biochemical properties may make these staphylococcal lipases isolated from facial sebaceous skin suitable for use as catalysts in the cosmetic, medicinal, food, or detergent industries.
ESTHER : Xie_2012_J.Microbiol.Biotechnol_22_84
PubMedSearch : Xie_2012_J.Microbiol.Biotechnol_22_84
PubMedID: 22297223

Title : Transesterification using the cross-linked enzyme aggregate of Photobacterium lipolyticum lipase M37 - Han_2011_J.Microbiol.Biotechnol_21_1159
Author(s) : Han JY , Kim HK
Ref : J Microbiol Biotechnol , 21 :1159 , 2011
Abstract : Biodiesel is methyl and ethyl esters of long-chain fatty acids produced from vegetable oils or animal fats. Lipase enzymes have occasionally been used for the production of this biofuel. Recently, biodiesel production using immobilized lipase has received increased attention. Through enhanced stability and reusability, immobilized lipase can contribute to the reduction of the costs inherent to biodiesel production. In this study, methanol-tolerant lipase M37 from Photobacterium lipolyticum was immobilized using the cross-linked enzyme aggregate (CLEA) method. Lipase M37 has a high lysine content (9.7%) in its protein sequence. Most lysine residues are located evenly over the surface of the protein, except for the lid structure region, which makes the CLEA preparation yield quite high (~93%). CLEA M37 evidences an optimal temperature of 30oC, and an optimal pH of 9-10. It was stable up to 50 degrees C and in a pH range of 4.0-11.0. Both soluble M37 and CLEA M37 were stable in the presence of high concentrations of methanol, ethanol, 1-propanol, and nbutanol. That is, their activities were maintained at solvent concentrations above 10% (v/v). CLEA M37 could produce biodiesel from olive oil and alcohols such as methanol and ethanol. Additionally, CLEA M37 generated biodiesel via both 2-step methanol feeding procedures. Considering its physical stability and reusability, CLEA M37 may potentially be used as a catalyst in organic synthesis, including the biodiesel production reaction.
ESTHER : Han_2011_J.Microbiol.Biotechnol_21_1159
PubMedSearch : Han_2011_J.Microbiol.Biotechnol_21_1159
PubMedID: 22127127

Title : A novel cold-adapted esterase from Salinisphaera sp. P7-4: gene cloning, overproduction, and characterization - Kim_2011_J.Gen.Appl.Microbiol_57_357
Author(s) : Kim YO , Park IS , Kim HK , Nam BH , Jeong Kong H , Kim WJ , Kim DG , Kim KK , Lee SJ
Ref : J Gen Appl Microbiol , 57 :357 , 2011
Abstract : Salinisphaera sp. P7-4 was isolated from the intestine of silver whiting, Sillago japonicas caught in the Pacific Ocean, and the esterase gene was cloned using the shotgun method. The amino acid sequence deduced from the nucleotide sequence (951 bp) corresponded to a protein of 316 amino acid residues with a molecular weight of 34,443. The esterase had 46 and 44% identities with the esterase enzymes of Ralstonia eutropha JMP134 and Rhodopseudomonas palustris HaA2, respectively. The primary structure of P7-4 esterase showed the conserved catalytic triad (Ser, Asp, His), consensus pentapeptide GXSXG, and oxyanion hole sequence (HG). The protein P7-4 was successfully expressed in Escherichia coli in a biologically active form. The enzyme showed high catalytic activity at low temperatures (5-25 degrees C) with an activation energy of 2.18 kcal/mol. This result indicated that the esterase from Salinisphaera sp. P7-4 is a new cold-adapted enzyme. The enzyme preferentially hydrolyzed acyl-group chains with short chain lengths of </=10 carbon. Metal ions such as Cd2(+), Co2(+), Cu2(+), Hg2(+), Ni2(+) and Zn2(+) inhibited enzymatic activity. Additionally, EDTA has no effect on its activity, whereas inhibition was observed with PMSF, a serine hydrolase inhibitor.
ESTHER : Kim_2011_J.Gen.Appl.Microbiol_57_357
PubMedSearch : Kim_2011_J.Gen.Appl.Microbiol_57_357
PubMedID: 22353741
Gene_locus related to this paper: 9gamm-h8ypm9

Title : Effects of methoxsalen from Poncirus trifoliata on acetylcholinesterase and trimethyltin-induced learning and memory impairment - Kim_2011_Biosci.Biotechnol.Biochem_75_1984
Author(s) : Kim JK , Choi SJ , Bae H , Kim CR , Cho HY , Kim YJ , Lim ST , Kim CJ , Kim HK , Peterson S , Shin DH
Ref : Biosci Biotechnol Biochem , 75 :1984 , 2011
Abstract : Previously, we identified methoxsalen (8-methoxy-2',3',6,7-furocoumarin) as the bioactive compound probably responsible for acetylcholinesterase (AchE) inhibition achieved by feeding crude extract of Poncirus trifoliate. To confirm the activity of methoxsalen, Institute of Cancer Research (ICR) mice were fed a control or a methoxsalen-supplemented diet for 4 weeks, and then learning and memory enhancing effects with respect to trimethyltin (TMT)-induced neurotoxicity were evaluated. The brain tissues of ICR mice were dissected after completion of the behavioral tests for biochemical analysis. Methoxsalen effectively reversed TMT-induced memory impairment on both Y-maze and passive avoidance tests. Brain AchE activity was inhibited by the oral consumption of all concentrations of methoxsalen. Moreover, the level of oxidative stress was significantly ameliorated in the groups on methodsalen containing diets. This is the first in vivo study conducted with methoxsalen in the field of AD research, and it indicates that further investigation of methoxsalen is warranted.
ESTHER : Kim_2011_Biosci.Biotechnol.Biochem_75_1984
PubMedSearch : Kim_2011_Biosci.Biotechnol.Biochem_75_1984
PubMedID: 21979070

Title : Neostigmine for the treatment of acute colonic pseudo-obstruction (ACPO) in pediatric hematologic malignancies - Lee_2010_Korean.J.Hematol_45_62
Author(s) : Lee JW , Bang KW , Jang PS , Chung NG , Cho B , Jeong DC , Kim HK , Im SA , Lim GY
Ref : Korean J Hematol , 45 :62 , 2010
Abstract : BACKGROUND: Acute colonic pseudo-obstruction (ACPO) refers to dilatation of the colon and decreased bowel motility without evidence of mechanical obstruction. Neostigmine, an acetylcholinesterase inhibitor, has been used in patients in whom supportive therapy failed to resolve ACPO. Here, we report the results of administering neostigmine to treat ACPO in children with hematologic malignancies. METHODS: Between September 2005 and December 2009, 10 patients (8 male and 2 female) were diagnosed with ACPO at the Department of Pediatrics, Catholic University of Korea. Diagnosis of ACPO was based on typical clinical features as well as colonic dilatation found on abdominal CT imaging. Neostigmine was administered subcutaneously at a dosage of 0.01 mg/kg/dose (maximum 0.5 mg) twice daily for a maximum of 5 total doses. ACPO was determined to be responsive to neostigmine if the patient showed both stool passage and improvement of clinical symptoms. RESULTS: The study group included 8 acute lymphoblastic leukemia patients, 1 patient with malignant lymphoma, and 1 patient with juvenile myelomonocytic leukemia. The median age at ACPO diagnosis was 8.5 years (range, 3-14). Overall, 8 patients (80%) showed therapeutic response to neostigmine at a median of 29 hours after the initial administration (range, 1-70). Two patients (20%) showed side effects of grade 2 or above, but none complained of cardiovascular symptoms that required treatment. CONCLUSION: In this study, ACPO was diagnosed most often in late-childhood ALL patients. Subcutaneous neostigmine can be used to effectively treat ACPO diagnosed in children with hematologic malignancies without major cardiovascular complications.
ESTHER : Lee_2010_Korean.J.Hematol_45_62
PubMedSearch : Lee_2010_Korean.J.Hematol_45_62
PubMedID: 21120165

Title : Ameliorative effect of 1,2-benzenedicarboxylic acid dinonyl ester against amyloid beta peptide-induced neurotoxicity - Jung_2009_Amyloid_16_15
Author(s) : Jung Choi S , Kim MJ , Jin Heo H , Kim JK , Jin Jun W , Kim HK , Kim EK , Ok Kim M , Yon Cho H , Hwang HJ , Jun Kim Y , Shin DH
Ref : Amyloid , 16 :15 , 2009
Abstract : Amyloid beta peptide (Abeta)-induced oxidative stress may be linked to neurodegenerative disease. Ethanol extracts of Rosa laevigata protected PC12 cells from hydrogen peroxide-induced oxidative stress. (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) reduction assays revealed a significant increase in cell viability when oxidatively stressed PC12 cells were treated with R. laevigata extract. The effect of R. laevigata on oxidative stress-induced cell death was further investigated by lactate dehydrogenase release assays and trypan blue exclusion assays. Administration of 1,2-benzenedicarboxylic acid dinonyl ester from R. laevigata extract to mice infused with Abeta significantly reversed learning and memory impairment in behavioural tests. After behavioural testing, the mice were sacrificed and brains were collected for the examination of lipid peroxidation, catalase activity and acetylcholinesterase (AchE) activity. These results suggest that 1,2-benzenedicarboxylic acid dinonyl ester from R. laevigata extract may be able to reduce Abeta-induced neurotoxicity, possibly by reducing oxidative stress. Therefore, R. laevigata extract may be useful for the prevention of oxidative stress-induced neurodegenerative disorders.
ESTHER : Jung_2009_Amyloid_16_15
PubMedSearch : Jung_2009_Amyloid_16_15
PubMedID: 19291510

Title : Cloning and characterization of a new cold-active lipase from a deep-sea sediment metagenome - Jeon_2009_Appl.Microbiol.Biotechnol_81_865
Author(s) : Jeon JH , Kim JT , Kim YJ , Kim HK , Lee HS , Kang SG , Kim SJ , Lee JH
Ref : Applied Microbiology & Biotechnology , 81 :865 , 2009
Abstract : To search for new cold-active lipases, a metagenomic library was constructed using cold-sea sediment samples at Edison Seamount and was screened for lipolytic activities by plating on a tricaprylin medium. Subsequently, a fosmid clone was selected, and the whole sequence of 36 kb insert of the fosmid clone was determined by shotgun sequencing. The sequence analysis revealed the presence of 25 open reading frames (ORF), and ORF20 (EML1) showed similarities to lipases. Phylogenetic analysis of EML1 suggested that the protein belonged to a new family of esterase/lipase together with LipG. The EML1 gene was expressed in Escherichia coli, and purified by metal-chelating chromatography. The optimum activity of the purified EML1 (rEML1) occurred at pH 8.0 and 25 degrees C, respectively, and rEML1 displayed more than 50% activity at 5 degrees C. The activation energy for the hydrolysis of olive oil was determined to be 3.28 kcal/mol, indicating that EML1 is a cold-active lipase. rEML1 preferentially hydrolyzed triacylglycerols acyl-group chains with long chain lengths of > or = 8 carbon atoms and displayed hydrolyzing activities toward various natural oil substrates. rEML1 was resistant to various detergents such as Triton X-100 and Tween 80. This study represents an example which developed a new cold-active lipase from a deep-sea sediment metagenome.
ESTHER : Jeon_2009_Appl.Microbiol.Biotechnol_81_865
PubMedSearch : Jeon_2009_Appl.Microbiol.Biotechnol_81_865
PubMedID: 18773201
Gene_locus related to this paper: 9bact-a0ejl0

Title : Inhibitory effect of Poncirus trifoliate on acetylcholinesterase and attenuating activity against trimethyltin-induced learning and memory impairment - Kim_2009_Biosci.Biotechnol.Biochem_73_1105
Author(s) : Kim JK , Bae H , Kim MJ , Choi SJ , Cho HY , Hwang HJ , Kim YJ , Lim ST , Kim EK , Kim HK , Kim BY , Shin DH
Ref : Biosci Biotechnol Biochem , 73 :1105 , 2009
Abstract : Various native Korean plants were screened to find an effective acetylcholinesterase (AChE) inhibitor for the treatment of Alzheimer's disease (AD). Among these plants, the ethanol extract of Poncirus trifoliate was selected for isolating the AChE inhibitor because it exhibited the highest inhibitory activity (47.31%). To separate the active compound from Poncirus trifoliate, solvent partition, open column chromatography, thin-layer chromatography (TLC), and high-performance liquid chromatography (HPLC) were utilized. The putative chemical structure of the AChE inhibitor was identified as methoxsalen by successive analysis with electron ionization mass spectrometry (EI-MS) and (13)C/(1)H-nuclear magnetic resonance (NMR). To confirm the attenuating effect of the Poncirus trifoliate extract against trimethyltin (TMT)-induced neurotoxicity, in vivo behavior tests were carried out. Our findings suggest that the Poncirus trifoliate extract significantly reversed TMT-induced learning and memory impairment. These results demonstrate that the Poncirus trifoliate extract could possess a wide range of beneficial activities for neurodegenerative disorders, notably AD.
ESTHER : Kim_2009_Biosci.Biotechnol.Biochem_73_1105
PubMedSearch : Kim_2009_Biosci.Biotechnol.Biochem_73_1105
PubMedID: 19420715

Title : In crystallo capture of a Michaelis complex and product-binding modes of a bacterial phosphotriesterase - Jackson_2008_J.Mol.Biol_375_1189
Author(s) : Jackson CJ , Foo JL , Kim HK , Carr PD , Liu JW , Salem G , Ollis DL
Ref : Journal of Molecular Biology , 375 :1189 , 2008
Abstract : The mechanism by which the binuclear metallophosphotriesterases (PTEs, E.C. catalyse substrate hydrolysis has been extensively studied. The mu-hydroxo bridge between the metal ions has been proposed to be the initiating nucleophile in the hydrolytic reaction. In contrast, analysis of some biomimetic systems has indicated that mu-hydroxo bridges are often not themselves nucleophiles, but act as general bases for freely exchangeable nucleophilic water molecules. Herein, we present crystallographic analyses of a bacterial PTE from Agrobacterium radiobacter, OpdA, capturing the enzyme-substrate complex during hydrolysis. This model of the Michaelis complex suggests the alignment of the substrate will favour attack from a solvent molecule terminally coordinated to the alpha-metal ion. The bridging of both metal ions by the product, without disruption of the mu-hydroxo bridge, is also consistent with nucleophilic attack occurring from the terminal position. When phosphodiesters are soaked into crystals of OpdA, they coordinate bidentately to the beta-metal ion, displacing the mu-hydroxo bridge. Thus, alternative product-binding modes exist for the PTEs, and it is the bridging mode that appears to result from phosphotriester hydrolysis. Kinetic analysis of the PTE and promiscuous phosphodiesterase activities confirms that the presence of a mu-hydroxo bridge during phosphotriester hydrolysis is correlated with a lower pK(a) for the nucleophile, consistent with a general base function during catalysis.
ESTHER : Jackson_2008_J.Mol.Biol_375_1189
PubMedSearch : Jackson_2008_J.Mol.Biol_375_1189
PubMedID: 18082180

Title : A new stilbene glucoside gallate from the roots of Polygonum multiflorum - Kim_2008_Arch.Pharm.Res_31_1225
Author(s) : Kim HK , Choi YH , Choi JS , Choi SU , Kim YS , Lee KR , Kim YK , Ryu SY
Ref : Arch Pharm Res , 31 :1225 , 2008
Abstract : A new stilbenoid (1) was isolated from the root extract of Polygonum multiflorum together with eight known constituents (2-9). The chemical structure of 1 was established as the 6''-O-monogalloyl ester of (E)-2,3,4',5-beta-tetrahydroxystilbene-2-beta-D-glucopyranoside based on physicochemical and spectroscopic analyses, particularly by NMR spectroscopic data, i.e., COSY, HMQC and HMBC. Compound 1 weakly inhibited acetylcholinesterase in vitro.
ESTHER : Kim_2008_Arch.Pharm.Res_31_1225
PubMedSearch : Kim_2008_Arch.Pharm.Res_31_1225
PubMedID: 18958411

Title : Structural basis for the cold adaptation of psychrophilic M37 lipase from Photobacterium lipolyticum - Jung_2008_Proteins_71_476
Author(s) : Jung SK , Jeong DG , Lee MS , Lee JK , Kim HK , Ryu SE , Park BC , Kim JH , Kim SJ
Ref : Proteins , 71 :476 , 2008
Abstract : The M37 lipase from Photobacterium lipolyticum shows an extremely low activation energy and strong activity at low temperatures, with optimum activity seen at 298 K and more than 75% of the optimum activity retained down to 278 K. Though the M37 lipase is most closely related to the filamentous fungal lipase, Rhizomucor miehei lipase (RML) at the primary structure level, their activity characteristics are completely different. In an effort to identify structural components of cold adaptation in lipases, we determined the crystal structure of the M37 lipase at 2.2 A resolution and compared it to that of nonadapted RML. Structural analysis revealed that M37 lipase adopted a folding pattern similar to that observed for other lipase structures. However, comparison with RML revealed that the region beneath the lid of the M37 lipase included a significant and unique cavity that would be occupied by a lid helix upon substrate binding. In addition, the oxyanion hole was much wider in M37 lipase than RML. We propose that these distinct structural characteristics of M37 lipase may facilitate the lateral movement of the helical lid and subsequent substrate hydrolysis, which might explain its low activation energy and high activity at low temperatures.
ESTHER : Jung_2008_Proteins_71_476
PubMedSearch : Jung_2008_Proteins_71_476
PubMedID: 18186467
Gene_locus related to this paper: 9gamm-q5drn8

Title : Functional expression and refolding of new alkaline esterase, EM2L8 from deep-sea sediment metagenome - Park_2007_Protein.Expr.Purif_52_340
Author(s) : Park HJ , Jeon JH , Kang SG , Lee JH , Lee SA , Kim HK
Ref : Protein Expr Purif , 52 :340 , 2007
Abstract : A metagenomics approach is an efficient method of isolating novel and useful genes from uncultured microorganisms in diverse environments. In this research, a gene encoding a new esterase (EM2L8) was cloned and characterized from the metagenomic DNA library of a deep-sea sediment. The gene consisted of 804bp encoding a polypeptide of 267 amino acids with a molecular mass of 28,952. The deduced amino acid sequence showed similarities with the BioH of Kurthia, the 3-oxoadipate enol-lactonase of Haloarcula and the acyltransferase of Thermoanaerobacter, which feature identities of 38%, 32%, and 33%, respectively. Residues essential for esterase activity, such as pentapeptide (GXSXG) and catalytic triad sequences, were uncovered. While the protein was overproduced mainly as inclusion body at 37 degrees C, it was mainly produced as a soluble active enzyme at 18 degrees C. A zymogram analysis revealed that purified EM2L8 taken from the soluble fraction could hydrolyze tributyrin substrate. Furthermore, the protein from the inclusion body fraction also showed strong activity on gel, thus indicating that the protein was refolded during SDS-gel electrophoresis and the ensuing incubation period. When the inclusion body was mixed with some anionic detergent solutions and diluted with a non-detergent buffer, the insoluble EM2L8 refolded rapidly and recovered its full esterase activity. Although EM2L8 had an optimum temperature of 50-55 degrees C, its activation energy in the range of 10-40 degrees C was 8.34kcal/mol, indicating that it is a cold-adapted enzyme. Moreover, it was found to have an optimum pH of 10-11, thus revealing that it is an alkaline enzyme. In this paper, the new esterase EM2L8 buried in a deep-sea sediment became known on the surface and was characterized biochemically.
ESTHER : Park_2007_Protein.Expr.Purif_52_340
PubMedSearch : Park_2007_Protein.Expr.Purif_52_340
PubMedID: 17126562
Gene_locus related to this paper: 9zzzz-EM2L8

Title : New cold-adapted lipase from Photobacterium lipolyticum sp. nov. that is closely related to filamentous fungal lipases - Ryu_2006_Appl.Microbiol.Biotechnol_70_321
Author(s) : Ryu HS , Kim HK , Choi WC , Kim MH , Park SY , Han NS , Oh TK , Lee JK
Ref : Applied Microbiology & Biotechnology , 70 :321 , 2006
Abstract : A Photobacterium strain, M37, showing lipolytic activity, was previously isolated from an intertidal flat of the Yellow Sea in Korea and identified as Photobacterium lipolyticum sp. nov. In the present study, the corresponding gene was cloned using the shotgun method. The amino acid sequence deduced from the nucleotide sequence (1,023 bp) corresponded to a protein of 340 amino acid residues with a molecular weight of 38,026. No sequence similarity was found with any known bacterial lipases/esterases; instead, the most similar enzymes were several filamentous fungal lipases. Although the similarity was very low (less than 16%), there were many conserved regions over the entire sequence and N-terminal oxyanion hole (RG) region, a signature sequence of filamentous fungal lipases. The novel protein M37 was produced in both a soluble and insoluble form when the Escherichia coli cells harboring the gene were cultured at 18 degrees C. The soluble protein exhibited lipase activity in a pH-stat assay using an olive oil emulsion. The M37 lipase also displayed a maximum activity at 25 degrees C and maintained its activity at a low temperature range (5-25 degrees C) with an activation energy (E(a)) of 2.07 kcal/mol. Accordingly, these results indicate that the M37 lipase from P. lipolyticum sp. nov. is a new cold-adapted enzyme.
ESTHER : Ryu_2006_Appl.Microbiol.Biotechnol_70_321
PubMedSearch : Ryu_2006_Appl.Microbiol.Biotechnol_70_321
PubMedID: 16088345
Gene_locus related to this paper: 9gamm-q5drn8

Title : Following directed evolution with crystallography: structural changes observed in changing the substrate specificity of dienelactone hydrolase - Kim_2005_Acta.Crystallogr.D.Biol.Crystallogr_61_920
Author(s) : Kim HK , Liu JW , Carr PD , Ollis DL
Ref : Acta Crystallographica D Biol Crystallogr , 61 :920 , 2005
Abstract : The enzyme dienelactone hydrolase (DLH) has undergone directed evolution to produce a series of mutant proteins that have enhanced activity towards the non-physiological substrates alpha-naphthyl acetate and p-nitrophenyl acetate. In terms of steady-state kinetics, the mutations caused a drop in the K(m) for the hydrolysis reaction with these two substrates. For the best mutant, there was a 5.6-fold increase in k(cat)/K(m) for the hydrolysis of alpha-naphthyl acetate and a 3.6-fold increase was observed for p-nitrophenyl acetate. For alpha-naphthyl acetate the pre-steady-state kinetics revealed that the rate constant for the formation of the covalent intermediate had increased. The mutations responsible for the rate enhancements map to the active site. The structures of the starting and mutated proteins revealed small changes in the protein owing to the mutations, while the structures of the same proteins with an inhibitor co-crystallized in the active site indicated that the mutations caused significant changes in the way the mutated proteins recognized the substrates. Within the active site of the mutant proteins, the inhibitor was rotated by about 180 degrees with respect to the orientation found in the starting enzyme. This rotation of the inhibitor caused the displacement of a large section of a loop on one side of the active site. Residues that could stabilize the transition state for the reaction were identified.
ESTHER : Kim_2005_Acta.Crystallogr.D.Biol.Crystallogr_61_920
PubMedSearch : Kim_2005_Acta.Crystallogr.D.Biol.Crystallogr_61_920
PubMedID: 15983415
Gene_locus related to this paper: psepu-clcd1

Title : Occurrence of ofloxacin ester-hydrolyzing esterase from Bacillus niacini EM001. - Kim_2004_J.Mol.Catal.B.Enzym_27_237
Author(s) : Kim HK , Na HS , Park MS , Oh TK , Lee TS
Ref : J Mol Catal B Enzym , 27 :237 , 2004
Abstract : A Bacillus niacini strain (EM001) producing an ofloxacin ester-enantioselective esterase was isolated from the soil samples collected near Taejon, Korea. The cloned gene showed that the esterase EM001 composed of 495 amino acids corresponding to a relative molecular weight (Mr) of 54,098 kDa. Based on the Mr and the protein sequence, the esterase EM001 was similar to p-nitrobenzyl esterase from Bacillus subtilis with an identity of 41.8%. The optimum temperature and pH of the purified His-tagged enzyme were 45 degC and 9.0, respectively. The purified esterase EM001 hydrolyzed preferably (R)-ofloxacin propyl ester than (S)-form ester at the initial reaction phase with an eeP of 67% until the conversion rate become up to 35%.
ESTHER : Kim_2004_J.Mol.Catal.B.Enzym_27_237
PubMedSearch : Kim_2004_J.Mol.Catal.B.Enzym_27_237
Gene_locus related to this paper: bacni-Q6KG48

Title : A novel lipase\/chaperone pair from Ralstonia sp. M1: analysis of the folding interaction and evidence for gene loss in R. solanacearum - Quyen_2004_Mol.Genet.Genomics_272_538
Author(s) : Quyen DT , Nguyen TT , Le TT , Kim HK , Oh TK , Lee JK
Ref : Mol Genet Genomics , 272 :538 , 2004
Abstract : A microbial strain (referred to as M1) that produces an extracellular lipase was isolated from a soil sample in Vietnam, and identified as a Ralstonia species by partial sequencing of its 16S rDNA. A genomic library was constructed from Pst I fragments, and a colony showing lipase activity was selected for further analysis. Sequencing of the 4.7-kb insert in this clone (named M1-72) revealed one incomplete and three complete ORFs, predicted to encode a partial hypothetical glutaminyl tRNA synthetase (304 aa), a hypothetical transmembrane protein (500 aa), a lipase (328 aa) and a lipase chaperone (352 aa), respectively. Alignment of the insert sequence with the corresponding region of the genome of R. solanacearum GMI1000 (GenBank Accession No. AL646081) confirmed the presence in the latter of the genes for the hypothetical transmembrane protein and glutaminyl tRNA synthetase, which exhibited 89-91% identity to their counterparts in M1. However, R. solanacearum GMI1000 lacks the complete lipase-encoding gene and the major part of the chaperone-encoding gene, creating a so-called "black hole". The deduced amino acid sequences of the products of the lipase gene lipA and chaperone gene lipB from strain M1 shared 49.3-60.3% and 23.9-32.7% identity, respectively, with those of the Burkholderia lipase/chaperone subfamily I.2. lipB is located downstream of lipA, and separated from it by only 9 bp, and each gene has a putative ribosome binding site. The mature lipase LipA, a His-tagged derivative (LipAhis), the tagged full-length chaperone LipBhis and a truncated form (DeltaLipBhis) lacking the 56 N-terminal residues were expressed in Escherichia coli BL21. LipA, LipAhis and DeltaLipBhis could be expressed at high levels (70, 15 and 12 mg/g wet cells, respectively) and were easily purified. However, LipBhis was expressed at a much lower level which precluded purification. The specific activity of purified LipAhis, expressed on its own, was very low (<52 U/mg). However, after co-incubation with the purified DeltaLipBhis in vitro, the specific activity of the enzyme was markedly enhanced, indicating that the chaperone facilitated correct folding of the enzyme. A lipase:chaperone ratio of 1:10 was found to be optimal, yielding an enzyme preparation with a specific activity of 650 U/mg.
ESTHER : Quyen_2004_Mol.Genet.Genomics_272_538
PubMedSearch : Quyen_2004_Mol.Genet.Genomics_272_538
PubMedID: 15668771
Gene_locus related to this paper: 9rals-q67dx2

Title : Effects of green tea polyphenol on cognitive and acetylcholinesterase activities - Kim_2004_Biosci.Biotechnol.Biochem_68_1977
Author(s) : Kim HK , Kim M , Kim S , Chung JH
Ref : Biosci Biotechnol Biochem , 68 :1977 , 2004
Abstract : The effect of tea polyphenol (TP) on cognitive and anti-cholinesterase activity was examined in scopolamine-treated mice. Chronic administration of TP significantly reversed scopolamine-induced retention deficits in both step-through passive avoidance and spontaneous alternation behavior tasks. Furthermore, TP exhibited a dramatic inhibitory effect on acetylcholinesterase activity. This finding suggests that TP might be useful in the treatment of Alzheimer's disease.
ESTHER : Kim_2004_Biosci.Biotechnol.Biochem_68_1977
PubMedSearch : Kim_2004_Biosci.Biotechnol.Biochem_68_1977
PubMedID: 15388975

Title : Naringenin from Citrus junos has an inhibitory effect on acetylcholinesterase and a mitigating effect on amnesia - Heo_2004_Dement.Geriatr.Cogn.Disord_17_151
Author(s) : Heo HJ , Kim MJ , Lee JM , Choi SJ , Cho HY , Hong B , Kim HK , Kim E , Shin DH
Ref : Dementia & Geriatric Cognitive Disorders , 17 :151 , 2004
Abstract : This study was performed to identify safe and more effective acetylcholinesterase (AChE) inhibitors in the treatment of Alzheimer's disease. The total methanol extract of Citrus junos had a significant inhibitory effect on AChE in vitro. By sequential fractionation of C.junos, the active component was finally identified as naringenin. Naringenin inhibited AChE activity in a dose-dependent manner. In this study, we also evaluated the anti-amnesic activity of naringenin, a major flavanone constituent isolated from C. junos, in vivo using ICR mice with amnesia induced by scopolamine (1 mg/kg body weight). Naringenin, when administered to mice at 4.5 mg/kg body weight, significantly ameliorated scopolamine-induced amnesia as measured in both the passive avoidance and the Y-maze test. These results suggest that naringenin may be a useful chemopreventive agent against Alzheimer's disease.
ESTHER : Heo_2004_Dement.Geriatr.Cogn.Disord_17_151
PubMedSearch : Heo_2004_Dement.Geriatr.Cogn.Disord_17_151
PubMedID: 14739537

Title : Inhibitory effect of zeatin, isolated from Fiatoua villosa, on acetylcholinesterase activity from PC12 cells - Heo_2002_Mol.Cells_13_113
Author(s) : Heo HJ , Hong SC , Cho HY , Hong B , Kim HK , Kim EK , Shin DH
Ref : Mol Cells , 13 :113 , 2002
Abstract : Acetylcholinesterase (AChE) inhibitors, which enhance cholinergic transmission by reducing the enzymatic degradation of acetylcholine, are the only source of the compound that is currently approved for the treatment of Alzheimer's disease (AD). The methanol extract from Fiatoua villosa among 100 traditional edible plants that were tested, showed the most potent inhibitory effect (51%) on acetylcholinesterase in vitro. After the sequential solvent fractionation of the methanol extract of Fiatoua villosa, the active fraction was repeatedly subjected to open-column chromatography on silica gel. From the highest inhibitory fraction, the chloroform fraction (75%) on AChE, the single compound, was obtained by the Sep-Pak Cartridge (C18: reverse phase column). This compound was finally purified by HPLC (micro-bondapack C18 reverse phase column: 19 x 300 mm). According to the electron impact mass spectrometry (EI-MS), we confirmed that the molecular mass was 219 m/z. The structure of this compound was identified as zeatin [2-methyl-4-(1H-purine-6-ylamino)-2-buten-1-ol], one of the derivatives of purine adenine. The concentration that was required for 50% enzyme inhibition (IC50 value) was 1.09 x 10(-4) M. This study demonstrated that the zeatin from Fiatoua villosa appeared to be the most potent AChE inhibitor in AD.
ESTHER : Heo_2002_Mol.Cells_13_113
PubMedSearch : Heo_2002_Mol.Cells_13_113
PubMedID: 11911460

Title : Expression and characterization of Ca(2+)-independent lipase from Bacillus pumilus B26 - Kim_2002_Biochim.Biophys.Acta_1583_205
Author(s) : Kim HK , Choi HJ , Kim MH , Sohn CB , Oh TK
Ref : Biochimica & Biophysica Acta , 1583 :205 , 2002
Abstract : A lipase-producing Bacillus pumilus strain (B26) was isolated from a soil sample collected in Korea. The cloned gene showed that the lipase B26 composed of a 34-amino-acid signal sequence and a 181-amino-acid mature part corresponding to a molecular mass (M(r)) of 19,225. Based on the M(r) and the protein sequence, the lipase B26 belongs to the lipase family I.4. The optimum temperature and pH of the purified enzyme were 35 degrees C and 8.5, respectively. The lipase B26 showed a 'Ca(2+)-independent thermostability and catalytic activity'. These are novel properties observed for the first time in lipase B26 among all bacterial lipases and correspond with the suggestion that this enzyme had no Ca(2+)-binding motif around the catalytic His156 residue. This enzyme seems to be a true lipase based on the experimental results that it could hydrolyze various long-chain triglycerides (C(14)-C(18)) and triolein (C(18:1)) and that it showed a typical interfacial activation mechanism toward both tripropionin and p-nitrophenyl butyrate.
ESTHER : Kim_2002_Biochim.Biophys.Acta_1583_205
PubMedSearch : Kim_2002_Biochim.Biophys.Acta_1583_205
PubMedID: 12117564

Title : Novel zinc-binding center and a temperature switch in the Bacillus stearothermophilus L1 lipase - Jeong_2002_J.Biol.Chem_277_17041
Author(s) : Jeong ST , Kim HK , Kim SJ , Chi SW , Pan JG , Oh TK , Ryu SE
Ref : Journal of Biological Chemistry , 277 :17041 , 2002
Abstract : The bacterial thermoalkalophilic lipases optimally hydrolyze saturated fatty acids at elevated temperatures. They also have significant sequence homology with staphylococcal lipases, and both the thermoalkalophilic and staphylococcal lipases are grouped as the lipase family I.5. We report here the first crystal structure of the lipase family I.5, the structure of a thermoalkalophilic lipase from Bacillus stearothermophilus L1 (L1 lipase) determined at 2.0-A resolution. The structure is in a closed conformation, and the active site is buried under a long lid helix. Unexpectedly, the structure exhibits a zinc-binding site in an extra domain that accounts for the larger molecular size of the family I.5 enzymes in comparison to other microbial lipases. The zinc-coordinated extra domain makes tight interactions with the loop extended from the C terminus of the lid helix, suggesting that the activation of the family I.5 lipases may be regulated by the strength of the interactions. The unusually long lid helix makes strong hydrophobic interactions with its neighbors. The structural information together with previous biochemical observations indicate that the temperature-mediated lid opening is triggered by the thermal dissociation of the hydrophobic interactions.
ESTHER : Jeong_2002_J.Biol.Chem_277_17041
PubMedSearch : Jeong_2002_J.Biol.Chem_277_17041
PubMedID: 11859083
Gene_locus related to this paper: geost-lipas

Title : Inhibitory effect of ursolic acid purified from Origanum majorana L on the acetylcholinesterase - Chung_2001_Mol.Cells_11_137
Author(s) : Chung YK , Heo HJ , Kim EK , Kim HK , Huh TL , Lim Y , Kim SK , Shin DH
Ref : Mol Cells , 11 :137 , 2001
Abstract : We screened 139 herbal spices in search of the acetylcholinesterase (AChE) inhibitor from natural resources. AChE inhibitors, which enhance cholinergic transmission by reducing the enzymatic degradation of acetylcholine, are the only source of compound currently approved for the treatment of Alzheimer's Disease (AD). Among these herbs, edible plants and spices, the ethanol extract from Origanum majorana L. showed the highest inhibitory effect on AChE in vitro. By sequential fractionation of Origanum majorana L. the active component was finally identified as ursolic acid (3 beta-Hydroxyurs-12-en-28-oic acid). The ursolic acid of Origanum majorana L. inhibited AChE activity in a dose-dependent and competitive/non-competitive type. The Ki value (representing the affinity of the enzyme and inhibitor) of Origanum majorana L. ursolic acid was 6 pM, and that of tacrine was 0.4 nM. The concentration required for 50% enzyme inhibition of the active component (IC50 value) was 7.5 nM, and that of tacrine was 1 nM. This study demonstrated that the ursolic acid of Origanum majorana L. appeared to be a potent AChE inhibitor in Alzheimer's Disease.
ESTHER : Chung_2001_Mol.Cells_11_137
PubMedSearch : Chung_2001_Mol.Cells_11_137
PubMedID: 11355692

Title : Crystallization and preliminary X-ray analysis of a thermoalkalophilic lipase from Bacillus stearothermophilus L1 - Jeong_2001_Acta.Crystallogr.D.Biol.Crystallogr_57_1300
Author(s) : Jeong ST , Kim HK , Kim SJ , Pan JG , Oh TK , Ryu SE
Ref : Acta Crystallographica D Biol Crystallogr , 57 :1300 , 2001
Abstract : A thermoalkalophilic lipase from Bacillus stearothermophilus L1 (L1 lipase) was crystallized in two different crystal forms using a low concentration of the enzyme and a calcium-exchange process. The first, needle-like, crystal form, which diffracts to about 3.5 A, belongs to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 67.84, b = 72.96, c = 104.41 A. The second, monoclinic, crystal form, which behaves better than the first form for crystallographic analyses, belongs to the monoclinic space group C2 and has unit-cell parameters a = 119.62, b = 85.05, c = 98.36 A, beta = 99.73 degrees. From the monoclinic crystals, a native data set and a samarium-derivative data set were collected to 2.0 and 2.3 A resolution, respectively. The difference Patterson map between the two data sets shows strong heavy-atom peaks, indicating that the crystals are suitable for a high-resolution structure determination.
ESTHER : Jeong_2001_Acta.Crystallogr.D.Biol.Crystallogr_57_1300
PubMedSearch : Jeong_2001_Acta.Crystallogr.D.Biol.Crystallogr_57_1300
PubMedID: 11526325

Title : Inhibitory effect of Artemisia asiatica alkaloids on acetylcholinesterase activity from rat PC12 cells - Heo_2000_Mol.Cells_10_253
Author(s) : Heo HJ , Yang HC , Cho HY , Hong B , Lim ST , Park HJ , Kim KH , Kim HK , Shin DH
Ref : Mol Cells , 10 :253 , 2000
Abstract : We screened 42 Korean traditional tea plants to determine the inhibitory effect of acetylcholinesterase and attenuation of toxicity induced by amyloid-beta peptide, which were related to the treatment of Alzheimer's disease (AD). The methanolic extract from Artemisia asiatica among tested 42 tea plants, showed the highest inhibitory effect (48%) on acetylcholinesterase in vitro. The methanolic extract was further separated with n-hexane, chloroform, and ethyl acetate of water, in order. The chloroform solubles, which were high in inhibitory effect of acetylcholinesterase, were repeatedly subjected to open column chromatography on silica gel. From the highest inhibitory fraction (78%) on acetylcholinesterase, the single compound was obtained by the Sep-Pak Cartridge (C18: reverse phase column). This compound was found to react positively on Dragendorff's reagent (potassium bismuth iodide), which typically reacted with the alkaloid. This compound was purified by HPLC (mu-bondapack C18 reverse phase column: 3.9 x 150 mm). The IC50 (the concentration of 50% enzyme inhibition) value of this compound was 23 micrograms/ml and the inhibitory pattern on acetylcholinesterase was mixed with competitive/non-competitive type. We examined the effects of this compound on toxicity induced by A beta (25-35) in rat pheochromocytoma PC12 cells. Pretreatment of the PC12 cells for 2 h with an alkaloid of Artemisia asiatica (1200 microg/ml) reduced the toxicity induced by A beta. This study demonstrated that an alkaloid of Artemisia asiatica, which was metabolized to small molecule in digestive tract and then could pass through the blood-brain barrier, appeared to be an acetylcholinesterase inhibitor with a blocker of neurotoxicity induced by A beta in human brain causing Alzheimer's disease.
ESTHER : Heo_2000_Mol.Cells_10_253
PubMedSearch : Heo_2000_Mol.Cells_10_253
PubMedID: 10901162

Title : Gene cloning and characterization of thermostable lipase from Bacillus stearothermophilus L1 - Kim_1998_Biosci.Biotechnol.Biochem_62_66
Author(s) : Kim HK , Park SY , Lee JK , Oh TK
Ref : Biosci Biotechnol Biochem , 62 :66 , 1998
Abstract : The gene coding for an extracellular lipase of Bacillus stearothermophilus L1 was cloned in Escherichia coli. Sequence analysis showed an open reading frame of 1254 bp, which encodes a polypeptide of 417 amino acid residues. The polypeptide was composed of a signal sequence (29 amino acids) and a mature protein of 388 amino acids. An alanine replaces the first glycine in the conserved pentapeptide (Gly-X-Ser-X-Gly) around the active site serine. The expressed lipase was purified by hydrophobic interaction and ion exchange chromatography using buffers containing 0.02% (v/v) Triton X-100. The lipase was most active at 60-65 degrees C and in alkaline conditions around pH 9-10. The lipase had highest activity toward p-nitrophenyl caprylate among the synthetic substrates and tripropionin among the triglycerides. It hydrolyzed beef tallow and palm oil more rapidly than olive oil at 50 degrees C.
ESTHER : Kim_1998_Biosci.Biotechnol.Biochem_62_66
PubMedSearch : Kim_1998_Biosci.Biotechnol.Biochem_62_66
PubMedID: 9501519
Gene_locus related to this paper: geost-lipas

Title : Characterization of an alkaline lipase from Proteus vulgaris K80 and the DNA sequence of the encoding gene - Kim_1996_FEMS.Microbiol.Lett_135_117
Author(s) : Kim HK , Lee JK , Kim H , Oh TK
Ref : FEMS Microbiology Letters , 135 :117 , 1996
Abstract : A facultatively anaerobic bacterium producing an extracellular alkaline lipase was isolated from the soil collected near a sewage disposal plant in Korea and identified to be a strain of Proteus vulgaris. The molecular mass of the purified lipase K80 was estimated to be 31 kDa by SDS-PAGE. It was found to be an alkaline enzyme having maximum hydrolytic activity at pH 10, while fairly stable in a wide pH range from 5 to 11. The gene for lipase K80 was cloned in Escherichia coli. Sequence analysis showed an open reading frame of 861 bp coding for a polypeptide of 287 amino acid residues. The deduced amino acid sequence of the lipase gene had 46.3% identity to the lipase from Pseudomonas fragi.
ESTHER : Kim_1996_FEMS.Microbiol.Lett_135_117
PubMedSearch : Kim_1996_FEMS.Microbiol.Lett_135_117
PubMedID: 8598267
Gene_locus related to this paper: provu-lipas