Search

Female menopause is a hormone deficiency phenomenon that causes hot flashes, vaginal dryness, depression, nervous tension, insomnia, obesity, and bone loss. There are various hormone replacement therapy (HRT)-based menopausal treatments, but they are accompanied by side effects such as endometrial cancer and hyperplasia. To confirm the menopausal improvement effect of Polygonatum sibiricum (PS), we prepared an ovariectomized animal model, administered 17β-estradiol (E2) and PS, and analyzed various menopausal symptoms. PS restored vaginal epithelium thickness, by increasing the expression of estrogen receptors ERα (ESR1) and ERβ (ESR2), and increased serotonin concentration by reducing serotonin receptor 1 A (5-HT1A) and glucocorticoid receptor (Gr) expression. In addition, PS suppressed obesity by increasing HDL-C and decreasing LDL-C levels and improved the osteoporosis induced by ovariectomy. In particular, by controlling Hand2, Fgf2, and Faf9 expression through PR, the antiproliferative signal was suppressed in uterine epithelium, thereby reducing the risk of side effects of the administration of E2 alone. These results demonstrate that PS alleviates menopausal symptoms without causing endometrial hyperplasia.

Agmatine, involved in various modulatory actions in cellular mechanisms, is produced from arginine (Arg) by decarboxylation reaction using arginine decarboxylase (ADC, EC 4.1.1.19). The major obstacle of using wild-type Escherichia coli ADC (ADCes) in agmatine production is its sharp activity loss and instability at alkaline pH. Here, to overcome this problem, a new disulfide bond was rationally introduced in the decameric interface region of the enzyme. Among the mutants generated, W16C/D43C increased both thermostability and activity. The half-life (T1/2) of W16C/D43C at pH 8.0 and 60°C was 560 min, which was 280-fold longer than that of the wild-type, and the specific activity at pH 8.0 also increased 2.1-fold. Site-saturation mutagenesis was subsequently performed at the active site residues of ADCes using the disulfide-bond mutant (W16C/D43C) as a template. The best variant W16C/D43C/I258A displayed a 4.4-fold increase in the catalytic efficiency when compared with the wild-type. The final mutant (W16C/D43C/I258A) was successfully applied to in vitro synthesis of agmatine with an improved yield and productivity (>89.0% yield based on 100 mM of Arg within 5 h).

We propose to use cRFP (common Repository of FBS Proteins) in the MS (mass spectrometry) raw data search of cell secretomes. cRFP is a small supplementary sequence list of highly abundant fetal bovine serum proteins added to the reference database in use. The aim behind using cRFP is to prevent the contaminant FBS proteins from being misidentified as other proteins in the reference database, just as we would use cRAP (common Repository of Adventitious Proteins) to prevent contaminant proteins present either by accident or through unavoidable contacts from being misidentified as other proteins. We expect it to be widely used in experiments where the proteins are obtained from serum-free media after thorough washing of the cells, or from a complex media such as SILAC, or from extracellular vesicles directly.

In addition to the traditional method of vaccine development, the mRNA coronavirus vaccine, which is attractive as a challenging vaccination, recently opened a new era in vaccinology. Here we describe the EG-COVID which is a novel liposome-based mRNA candidate vaccine that encodes the spike (S) protein of SARS-CoV-2 with 2P-3Q substitution in European variant. We developed the mRNA vaccine platform that can be lyophilized using liposome-based technology. Intramuscular injection of the EG-COVID elicited robust humoral and cellular immune response to SARS-CoV-2. Furthermore, sera obtained from mice successfully inhibited SARS-CoV-2 viral infection into Vero cells. We developed EG-COVID and found it to be effective based on in vitro data, and we plan to initiate a clinical trial soon. Since EG-COVID is a lyophilized mRNA vaccine that is convenient for transportation and storage, accessibility to vaccines will be significantly improved.

A key point of protein stability engineering is to identify specific target residues whose mutations can stabilize the protein structure without negatively affecting the function or activity of the protein. Here, we propose a method called RiSLnet (Rapid identification of Smart mutant Library using residue network) to identify such residues by combining network analysis for protein residue interactions, identification of conserved residues, and evaluation of relative solvent accessibility. To validate its performance, the method was applied to four proteins, that is, T4 lysozyme, ribonuclease H, barnase, and cold shock protein B. Our method predicted beneficial mutations in thermal stability with ~62% average accuracy when the thermal stability of the mutants was compared with the ones in the Protherm database. It was further applied to lysine decarboxylase (CadA) to experimentally confirm its accuracy and effectiveness. RiSLnet identified mutations increasing the thermal stability of CadA with the accuracy of ~60% and significantly reduced the number of candidate residues (~99%) for mutation. Finally, combinatorial mutations designed by RiSLnet and in silico saturation mutagenesis yielded a thermally stable triple mutant with the half-life (T 1/2 ) of 114.9 min at 58°C, which is approximately twofold higher than that of the wild-type.

PURPOSE This study aimed to investigate whether hydrogen (H2) water drinking reduced oxidative damage products (8-hydroxy-2-deoxyguanosine [8-oxo-dG], malondialdehyde [MDA], L-lactate) produced in blood after exercise-induced oxidative stress. METHODS Men (n=7) and women (n=8) (systolic blood pressure [BP]: 90-150 mmHg, diastolic BP: 60-100 mmHg) were recruited. Exercise was performed for 20 minutes (8 km/hr) using treadmill. Placebo or test group drunk water and hydrogen water (1.2-1.5 ppm, 1,000 mL), respectively, right after exercise. Three days after, the same experiment was performed in the cross-over manner. Blood samples were obtained 30 minutes before exercise, right after exercise, and 30, 60, 120 minutes after exercise. 8-oxo-dG, MDA and L-lactate in serum were determined using ELISA kit. RESULTS The blood levels of 8-oxo-dG were significantly increased after exercise (from 381±9.4 to 524±14.9 ng/mL for male; from 367±13.5 to 509±10.7 ng/mL for female). In the placebo group, the increased 8-oxo-dG at each time was not significantly different from the peak level induced by the exercise except that at 120 minutes in female. In the test group, its levels at each time were significantly lower than that in the peak level. The effect of hydrogen water was more pronounced in female than in male. The results obtained with MDA were quite similar to those with 8-oxo-dG. The L-lactate levels were not significant statistically in both groups. CONCLUSIONS The data suggest that the one time drinking of hydrogen water may suppress the exercise-induced oxidative stress.

Twenty analogs of [Orn6,D-Ala9]α-factor were synthesized and assayed for their biological activities: seven analogs of [Orn6,X9]α-factor, seven analogs of [X6,D-Ala9]α-factor, five analogs of [X5,X6,D-Ala9]α-factor, and native α-factor (X = amino acids). Their biological activities (halo, gene induction, and affinity) were measured using S. cerevisiae Y7925 and LM102 and compared with those of native α-factor (100%). G protein-coupled receptor was expressed in strain LM102 containing pESC-LEU-STE2 vector. [Dap6,D-Ala9]α-factor with weak halo activity (10%) showed the highest receptor affinity (＞ 230%) and the highest gene induction activity (167%). [Arg6,D-Ala9]α-factor showed the highest halo activity (2,000%). The number of active binding sites per cell (about 20,000 for strain LM102) was determined using a newly-designed fluorescence-based detector, [Arg6,D-Ala9]α-factor-Edan, with high sensitivity (12,500-fold higher than the absorption-based detector [Orn6]α-factor-[Cys]3).

Tyrosinase is a type 3 copper oxygenase that catalyzes a phenol moiety into ortho-diphenol, and subsequently to ortho-quinone. Diverse tyrosinases have been observed across the kingdom including Animalia, Bacteria, Plantae, and Fungi. Among the tyrosinases, bacterial, and mushroom tyrosinases have been extensively exploited to prepare melanin, ortho-hydroxy-polyphenols, or novel plant secondary metabolites during the past decade. And their use as a biocatalyst to prepare various functional biocompounds have drawn great attention worldwide. Herein, we tailored a bacterial tyrosinase from Bacillus megaterium (BmTy) using circular permutation (CP) engineering technique which is a novel enzyme engineering technique to covalently link original N and C termini and create new termini on the middle of its polypeptide. To construct a smart rationally-designed CP library, we introduced 18 new termini at the edge of each nine loops that link α-helical secondary structure in BmTy. Among the small library, seven functional CP variants were successfully identified and they represented dramatic change in their enzyme characteristics including kinetic properties and substrate specificity. Especially, cp48, 102, and 245 showed dramatically decreased tyrosine hydroxylase activity, behaving like a catechol oxidase. Exploiting the dramatic increased polyphenol oxidation activity of cp48, orobol (3'-hydroxy-genistein) was quantitatively synthesized with 1.48 g/L, which was a 6-fold higher yield of truncated wild-type. We examined their kinetic characters through structural speculation, and suggest a strategy to solubilize the insoluble artificial variants effectively.

Ornithine decarboxylase (ODC) converts C5 ornithine into C4 putrescine, a monomer for polyamide synthesis. However, ODC also has minor activity towards cell metabolite C6 lysine and yields C5 cadaverine. The accumulation of cadaverine in the reaction solution causes increase in the operational cost of subsequent distillation process for putrescine purification. Here, to increase ODC substrate specificity toward ornithine over lysine, molecular modelling and protein network analysis, specifically k-clique community analysis, around the substrate tunnel were performed. This resulted in a mutant with two-fold increase in substrate specificity (ornithine versus lysine) without losing its original activity towards ornithine (kcat/KM = 61.5 s−1 mM-1), compared to the native enzyme. When this mutant was used for putrescine synthesis, 31.6 g/L putrescine (based on 51.5 g/L ornithine) titer was achieved, while 0.007 g/L (based on 2.57 g/L lysine) cadaverine was produced. This corresponds to four-fold decrease in cadaverine yield compared to the native ODC.

Thirteen analogs of tridecapeptide α-factor (WHWLQLKPGQPMY) of Saccharomyces cerevisiae with C- or N-terminal Trp extension and isosteric replacement by Aib at position 8 and 11, Trp at position 13, D-Ala at position 9, and Orn and Glu at position 6 were synthesized and assayed for their biological activity. Receptor binding assay was carried out using our newly developed spectrophotometric method with detector peptide 14. C- or N-terminal extended analogs, α-factor-[Trp]n (n =1-5) 1-5 and [N-Trp]1-α-factor 6, were all less active than native α-factor and gradual decreases in both activity and receptor affinity were observed with greater Trp extension. Trp-substituted analog at position 13, [Trp 13 ]α-factor 7, exhibited about 2-fold reductions in both activity and receptor affinity. Aib-substituted analogs, [Aib 8 ]α-factor 8 and [Aib 11 ]α-factor 9, showed 5- to 10fold reduction in activity as well as 3-fold reduction in receptor affinity compared to native α-factor. [Orn 6 ]αfactor 10 demonstrated strong potency with a 7.0-fold increase in halo activity as well as 1.8-fold increase in receptor affinity compared to native α-factor. For two double substituted analogs, [Glu 6 ,D-Ala 9 ]α-factor 12 showed the slightly decreased potency in halo activity compared to analog 10, whereas [Orn 6 ,D-Ala 9 ]α-factor 11 exhibited 15-fold higher halo activity as well as nearly 3-fold higher receptor affinity compared to native α-factor.

Background: Although ginsenosides such as Rg1, Rb1 and Rg3 have shown promise as potential nutraceuticals for cognitive impairment, their use has been limited due to high production cost and low potency. In particular, the process of extracting pure Rg3 from ginseng is laborious and expensive. Methods: We described the methods in preparing ginseol k-g3, an Rg3-enriched fraction, and evaluated its effects on scopolamine-induced memory impairment in mice. Results: Ginseol k-g3 (25–200 mg/kg) significantly reversed scopolamine-induced cognitive impairment in the passive avoidance, but not in Y-maze testing. Ginseol k-g3 (50 and 200 mg/kg) improved escape latency in training trials and increased swimming times within the target zone of the Morris water maze. The effect of ginseol k-g3 on the water maze task was more potent than that of Rg3 or Red ginseng. Acute or subchronic (6 d) treatment of ginseol k-g3 did not alter normal locomotor activity of mice in an open field. Ginseol k-g3 did not inhibit acetylcholinesterase activity, unlike donezepil, an acetylcholinesterase inhibitor. Rg3 enrichment through the ginseol k-g3 fraction enhanced the efficacy of Rg3 in scopolamine-induced memory impairment in mice as demonstrated in the Morris water maze task. Conclusion: The effects of ginseol k-g3 in ameliorating scopolamine-induced memory impairment in the passive avoidance and Morris water maze tests indicate its specific influence on reference or long-term memory. The mechanism underlying the reversal of scopolamine-induced amnesia by ginseol k-g3 is not yet known, but is not related to anticholinesterase-like activity.

Cirsium japonicum (CJ) has been shown to possess antidepressant-like properties. In the present study, we sought to identify which constituent of CJ might be responsible for its antidepressant effects and determine probable mechanism of action. The ethanol extract of CJ was administered to mice then behavioral changes were evaluated in the forced-swimming test (FST) and open-field test (OFT). In addition, its effects on norepinephrine (NE) reuptake and intracellular chloride (Cl(-)) flux were determined, in vitro. The effects of CJ's major constituents (linarin, pectolinarin, chlorogenic acid, luteolin) were also evaluated. CJ showed antidepressant-like effect by significantly reducing immobile behavior of mice in the FST, without increasing locomotor activity in the OFT. CJ had no effect on monoamine (NE) uptake, but it significantly promoted Cl(-) ion influx in human neuroblastoma cells. This CJ-induced Cl(-) influx was significantly blocked by co-administration of the competitive GABAA receptor antagonist, bicuculline. Among the major constituents of the CJ extract, only luteolin produced similar antidepressant-like effect, in vivo, and Cl(-) ion influx, in vitro. Altogether, the present results suggest that the antidepressant-like effect of CJ was most probably induced by its constituent luteolin, mediated through potentiation of the GABAA receptor-Cl(-) ion channel complex.

]α-factoramide ( 10) were found to have antagonistic activity. Analogs 3 and 6 showed greater antagonistic activity thananalogs 8 and 10. Key Words : α-Factor, Amide analogs, S. cerevisiae, ActivityIntroductionThe α-factor, tridecapeptide (WHWLQLKPGQPMY), issecreted by Saccharomyces cerevisiae MATa haploid cellsand recognized by a receptor that is coded by gene (STE2)which is expressed in S. cerevisiae MATa haploid cells.

본 연구는 한국 여성근로자의 직업관련 건강상태와 영향요인을 파악하기 위한 서술적 상관관계연구이다. 본 연구자료는 한국산업안전보건공단 한국산업안전보건연구원에서 취업자 근로환경 전국표본조사를 위해 2006년도 시행한 근로환경조사를 바탕으로 20-64세 여성근로자들로부터 얻었다. 한국 여성근로자의 직업관련 신체적 건강상태는 교육수준, 가정내 주수입원 종류, 회사규모, 고용형태, 주당 근로시간, 교대근무여부, 직무요구도 등에서 유의한 차이가 있었다. 심리적 건강상태는 교육수준, 월 평균소득, 가정내 주수입원 종류, 주당 근로시간, 교대근무 여부, 감정노동여부, 직무요구도, 직무자율성, 물리적 위험, 화학적 위험, 근골격계 위험부담요인 등에서 유의한 차이가 있었다. 직업관련 신체적 심리적 건강상태에 영향을 주는 요인을 알아보기 위해 단계적 중회귀분석을 한 결과 신체적 건강상태와 심리적 건강상태에 가장 큰 영향을 주는 요인은 근골격계 부담 이었고 설명력은 각각 6.12%, 2.97% 이었다. 그러므로 근골격계 부담요인 감소를 위한 작업장 환경조성과 체계적인 건강관리 및 보건교육이 필요할 것으로 사료된다. 【This is a descriptive correlation study to identify health status and affecting factors related to job among Korean women employees. Data were obtained from women employees aged 20 to 64 using the Korean Working Conditions Survey 2006(KWCS). Stepwise multiple regression analysis using SAS version 9.1 was performed to identify affecting factors on physical and psychological health status related to job among Korean women employees. Musculoskeletal work-load was the most significant factor affecting on both physical and psychological health status. Modification of work environment to relieve musculoskeletal work-load, systemic health management and health education needs to be given to women employees who were exposed to high musculoskeletal work-load.】

l -Tert-leucine (Tle) and l -3-hydroxyadamantylglycine of (HAG) are important intermediates for a variety of pharmaceutical classes. They were asymmetrically produced from corresponding keto acids using branched-chain aminotransferase (BCAT) with l -glutamate (Glu) as an amino donor. For the production of l -Tle and l -HAG, BCAT from Enterobacter sp. TL3 (BCATen) and BCAT from Escherichia coli K12 (ilvE, newly named as BCATes) were used, respectively. In our current study, we characterized the basic properties of BCATen and BCATes such as substrate specificity, enantioselectivity, and kinetic parameters. The activities of BCATen and BCATes were inhibited severely by α-ketoglutarate which is a deaminated product of l -Glu. In the presence of 10 mM α-ketoglutarate, both enzymes activities were reduced up to 80%. In order to overcome product inhibition by α-ketoglutarate and the problem of equilibrium of the transamination reaction, coupling reactions were carried out with l -glutamate dehydrogenase (GDH)/formate dehydrogenase (FDH) and AspAT. The coupling reaction dramatically increased the yields of both target compounds. 135 mM of l -Tle (>99% ee) was produced from 150 mM corresponding keto acid in BCATen/GDH/FDH coupling reaction with 90% conversion. In addition, 90.5 mM l -HAG (>99% ee) was produced from 100 mM corresponding keto acid in BCATes/AspAT coupling reaction using recombinant whole-cells.

A series of conjugated cyclic and linear enkephalin analogs, Tyr-c[D-A 2 bu-Gly-Phe-Asp(NH-X)], where X = methyl, stearyl or PEG 350 , and Tyr-D-Ala-Gly-Phe-Cys(S-X), where X = methyl, octyl, or farnesyl, were synthesized in solution to investigate the receptor selectivity of opioids based on Schwyzer's membrane compartment concepts. 5,6 Cyclizations of the target compounds were achieved in high yields (> 60%) employing BOP, NaHCO 3 in DMF despite the steric hindrance of the bulky pendant groups. In the binding assay, the hydrophobic fatty acyl conjugates retained μ-receptor selectivity. The unsaturated farnesyl conjugate exhibited the increased binding affinity than the saturated stearyl conjugate for both μ-and δ-opioid receptors. The PEG conjugates displayed the δ-receptor selectivity. The low molecular weight PEG 350 conjugate exhibited the increase selectivity than the high molecular weight PEG 5000 conjugate to the δ-receptor. The results of this study support the membrane compartment concepts.

An interlaboratory study was performed in eight laboratories to validate a liquid chromatography-tandem mass spectrometry (LC/MS/MS) method for the simultaneous determination of aflatoxins and sterigmatocystin (STC) in white rice and sorghum (Sorghum bicolor). Fortified samples (at three different levels) of white rice and sorghum were extracted, purified through a solid-phase extraction (SPE) column, and then analyzed by LC/MS/MS. The apparent recoveries (ARs) ranged from 78.8% to 95.0% for aflatoxins and from 85.3% to 96.7% for STC. The relative standard deviations for repeatability (RSDr) and reproducibility (RSDR) of aflatoxins were in the ranges 7.9%-33.8% and 24.4%-81.0%, respectively. For STC, the RSDr ranged from 7.1% to 40.2% and the RSDR ranged from 28.1% to 99.2%. The Horwitz ratio values for the aflatoxins and STC ranged from 0.4 to 1.2 in white rice and from 0.3 to 1.0 in sorghum, respectively. These results validated this method for the simultaneous determination of aflatoxins and STC by LC/MS/MS after SPE column cleanup. The percentages of satisfactory Z-score values (vertical bar Z vertical bar

A novel thermostable ω-transaminase from Thermomicrobium roseum which showed broad substrate specificity and high enantioselectivity was identified, expressed and biochemically characterized. The advantage of this enzyme to remove volatile inhibitory by-products was demonstrated by performing asymmetric synthesis and kinetic resolution at high temperature.

Because of Korea’s nationwide Internet capacity, information technology is accelerating the development of electronic democracy and the informatization of public administration in that country. This article analyzes how information technology affects organizational development, using the Korea Procurement Service as a case study. Based on Rockart, Earl, and Ross’s model, this study focuses on four aspects of redesign-operational redesign, support redesign, managerial redesign, and network process redesign. By exploring the link between information technology and other organizational factors, this article explores the effect of the introduction of an electronic system into this agency, which traditionally had a reputation for little corruption and red tape in Korea. Relying on previous studies and statistical data, this study explains the extraordinary high performance of the Korea Procurement Service and analyzes the effects of information technology on the achievement of service efficiency and corruption control.