• 한국미생물·생명공학회지
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• 제51권1호
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• pp.26-31
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• 2023

A map gene encoding methionyl-specific aminopeptidase (MAP; EC 3.4.11.18) was cloned from Tetragenococcus halophilus CY54. Translated amino acid sequence of CY54 MAP showed high similarities with those from Enterococcus faecalis (83.8%) and Streptococcus salivarius (62.2%) but low similarities with MAPs from Lactobacillus and Lactococcus genera. The map gene was overexpressed in E. coli BL21(DE3) using pET26b(+),pET26b(+), and the recombinant MAP was purified by using an Ni-NTA column. The size of recombinant MAP was 29 kDa as determined by SDS-PAGE. The optimum pH and temperature of CY54 MAP were pH 5.0 and 60℃, respectively. The activity of CY54 MAP was most significantly increased by Co²⁺ ion (159%), and showed the highest activity at 12% NaCl. K_m and V_max were 0.64 ± 0.006 mM and 10.12 ± 0.014 U/mg protein, respectively when met-pNA was used as the substrate. This is the first report on a MAP from Tetragenococcus species.

• Journal of Microbiology and Biotechnology
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• 제33권3호
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• pp.371-377
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• 2023

In this study, a pepA gene encoding glutamyl (aspartyl)-specific aminopeptidase (PepA; E.C. 3.4.11.7) was cloned from Tetragenococcus halophilus CY54. The translated PepA from T. halophilus CY54 showed very low similarities with PepAs from Lactobacillus and Lactococcus genera. The pepA from T. halophilus CY54 was overexpressed in E. coli BL21(DE3) using pET26b(+). The recombinant PepA was purified by using an Ni- NTA column. The size of the recombinant PepA was 39.13 kDa as determined by SDS-PAGE, while its optimum pH and temperature were pH 5.0 and 60℃, respectively. In addition, the PepA was completely inactivated by 1 mM EDTA, indicating its metallopeptidase nature. The Km and Vmax of the PepA were 0.98 ± 0.006 mM and 0.1 ± 0.002 mM/min, respectively, when Glu-pNA was used as the substrate. This is the first report on PepA from Tetragenococcus species.

• 대한의료기공학회지
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• 제15권1호
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• pp.61-70
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• 2015

Objectives : The purpose of this study is to investigate the effects of Su-Gi therapy for Insomnia by using ISI scores and PSQI test. Methods : We investigated 4 patients on condition of Insomnia who was hospitalized in the M Korean medicine hospital in Gyeonggi Province from 8th June, 2015 to 21st October, 2015. The Su-Gi therapy was treated on patients once a day for 10 minutes. We figured out the outcome by using ISI scores and PSQI test. Results : There were no differences on average sleeping time and the time to falling a sleep. Meaningful outcomes in values of ISI scores and PSQI test was not deduced after Su-Gi therapy. There were some progress on disturbance of daytime due to insomnia, ISI scores and PSQI test. In the Correlation analysis of outcomes of before and after the Su-Gi therapy, there were some improvements on ISI scores and PSQI tests and some questionnaire but were not enough to reach meaningful outcomes. We concluded that the more treatment time is needed to improve the condition of patients on insomnia as we see in PSQI test, one month is minimum period time to draw the conclusion. Conclusions : Theses outcomes suggest a possibilities that improvements who are suffering from insomnia can be get well when the Su-Gi therapy is performed for sufficient moments.

• 한국독어학회지:독어학
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• 제9집
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• pp.1-21
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• 2004

Die sogenannten verba pura bilden eine Teilmenge der germanischen starken Verben der VII. KLasse. Sie sind auf einsilbige langvokalisch auslautende Wurzeln des Vorurgermanischen $zur\"{u}ckzufuhren$: z. B. ${\ast}kn\bar{e}an$ 'erkennen', ${\ast}m\bar{e}an$ '$m\"{a}hen$', ${\ast}bl\bar{o}an$ $'bl\"{u}hen'$, ${\ast}m\bar{o}an$ $'qu\"{a}hlen'$. Um den Begriff der verba pura genau zu $erkl\"{a}ren$, stellt man diesen Verben die sogenannten verba impura $gegen\"{u}ber$, bei denen auf die lang Wurzelvokale urgerm. ${\ast}-\bar{e}-$ und ${\ast}-\ber{o}-$ ein Konsonant folgl: z.. B. urgerm. ${\ast}l\bar{e}tan$ (got. \etan) 'lassen', ${\ast}hw\bar{o}pan$ (got. hvopan) 'sich $r\"{u}hmen$'. Die germanischen starken Verben der VII. Klasse hat die $st\"{a}rksten$ Umgestaltungen mitgemacht, und zwar die verba pura noch mehr. In diesem Aufsatz untersuchte ich die $Gr\"{u}nde$ ihrer Umgestaltungen im germanischen Verbalsystem. Danach wird der Vorgang erschlossen, wie die verba pura in Ahd. zu den schwachen Verben I $\"{u}bertraten$. Dabei werden die folgenden Punkte besonders behandelt: Hatten die $fr\"{u}hzeitigen${\;}Pr\"{a}sensformen$ der verba pura -i-, das aus -jan herausgeht? Wenn nein, wie haben die verba pura zur schwachen Flexion $\"{u}bertraten$? Warum konnten die verba pura wie die anderen redupliziernden Verben in Ahd. nicht zu ablautenden Verben $\"{u}bergehen$?

• 한방안이비인후피부과학회지
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• 제21권1호
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• pp.70-82
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• 2008

Objective : The purpose of this study is to examine the effects of Baickbujasan(BB) on the skin damage and depigmentation. Method : The inhibition of tyrosinase activity, melanogenesis and cell viability in cultured B16 melanoma cells were measured. In order to test effects of reduction of melanogenesis, B16 F-10 mouse melanoma stem line was employed to extract melanin from cultured cell, where BB was added or not, and was dissolved in alkali for colorimetric analysis. Also, in order to test skin alteration in C57BL/6 after UV irradiation, the animals were grouped into a UV urradiation group and UV irradiation after BB application group. Dopa oxidase tissue staining was excuted to invesitage the change in the distribution of active melanin cell. The distribution of active melanin cell in inner skin of iNOS after damage from UVB irradiation and the manifestation condition of P53 which takes part in natural death of keratinocyte were examined. Result : The results indicate that BB has significant effects on tyrosinase activity, and melanogenesis in vivo test. BB seems to reduce C57BL/6, external dermatological damage, for instance, erythematous papule, eczema, loss of keratinocyte, reduction in pus, and relieves dermatological damages. Conclusion : BB can be applied externally for UV protection and depigmentation.

• 대한한방부인과학회지
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• 제21권4호
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• pp.1-16
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• 2008

Purpose: The purpose of this study is to investigate the effect of Boheo-tang (B) and Boheo-tang plus cervi pantotrichum cornu (B+CP) on lactation in postpartum C57BL/6N mice. Methods: Normal saline(control), Band B+CP (8mu l/g$) were administerd p.o. twice a day for 20 days. Lactating mammary gland tissues were examined through light microscope by the way of HE staining and immunohistochemical assay. Milk producing associated gene expression were accessed by RT-PCR. Results: In mammary gland, amount of adipose tissues were decreased in both Band B+CP treated group. And the ductal branches and alveolar tissues increased in both treated group. Immunoreactivity of prolactin receptors was increased both treated group, and immunoreactivity of oxytocin receptors was increased in the B+CP treated group. In both treated group, IGF-l mRNA expression was increased and TGF-$\beta$ mRNA expression was decreased. And PRL mRNA expression was increased in the B+CP treated group. PL-l mRNA expression was decreased in the B treated group but increased in the B+CP treated group. Conclusion: This study shows that treatment of Boheo-tang and Boheo-tang plus cervi pantotrichum cornu can improve postpartum lactation in C57BL/6N mice.

• Journal of Applied Biological Chemistry
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• 제53권4호
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• pp.195-201
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• 2010

B. pseudomycoides로 부터 alanine racemase 유전자를 분리한 다음 이 효소에 존재하는 두개의 cysteine을 하나(C316A) 또는 두개 모두(C316-365A) alanine으로 치환시켰다. 치환된 alanine racemamase는 pET-21 운반체에 삽입한 다음 숙주세포로서 E. coli BL21 (DE3)를 이용하여 발현시켰다. 발현된 단백질은 6XHis이 결합된 affinity chromatography를 이용하여 분리하였으며 SDS-PAGE 분석에서 모두 약 46 kDa의 주요 단일밴드를 나타내었다. Cysteine(-) 변이체의 alanine racemase가 모두 활성도를 보여 cysteine이 catalytic 또는 binding sit에 관여하지 않는 것으로 추정되었다. 변이체 효소들은 wild type에 비하여 열 안정성이 모두 떨어져 $60^{\circ}C$ pH 8.0에서의 활성도 반감시간이 각각 26(wild type), 21(C316A) 18분(C316-365A)-을 나타내었다. 이러한 결과는 cysteine이 열안정화에 상당히 기여함을 알 수 있었다. 그러나 pH 변화에 대한 안정성은 큰 차이가 없었다.

• 한국미생물·생명공학회지
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• 제42권2호
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• pp.190-193
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• 2014

CMP-Neu5Ac synthetase는 sialyated 된 glycoconjugates의 전구체로 사용되는 CMP-Neu5Ac를 합성하는데 관여하는 주요 효소이다. Escherichia coli K1에서 유래한 CMP-Neu5Ac synthetase 유전자 (neuA)는 평소 E. coli BL21(DE3)에서 비수용성으로 생성되는데, 이를 수용성 단백질로 생산하고자 여러 가지 샤페론 단백질 동시 발현기술을 이용하였다. 이를 위해, GroEL-ES와 DnaK-DnaJ-GrpE를 암호화하는 pG-KJE8 plasmid와 neuA를 동시 형질전환 시켰고 0.01 mM IPTG와 0.005 mg/ml의 L-arabinose로 유도하여 $20^{\circ}C$에서 발현시켰다. 그 결과, E. coli에서의 수용성 CMP-Neu5Ac Synthetase 생산이 현저하게 증가하였다.

• Journal of Microbiology and Biotechnology
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• 제25권5호
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• pp.696-703
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• 2015

A gamma-aminobutyric acid (GABA)-producing microorganism was isolated from jeot-gal (anchovy), a Korean fermented seafood. The isolate, A156, produced GABA profusely when incubated in MRS broth with monosodium glutamate (3% (w/v)) at 37℃ for 48 h. A156 was identified as Lactobacillus sakei by 16S rRNA gene sequencing. The GABA conversion yield was 86% as determined by GABase enzyme assay. The gadB gene encoding glutamate decarboxylase (GAD) was cloned by PCR. gadC encoding a glutamate/GABA antiporter was located immediately upstream of gadB. The operon structure of gadCB was confirmed by RT-PCR. gadB was overexpressed in Escherichia coli BL21(DE3) and recombinant GAD was purified. The purified GAD was 54.4 kDa in size by SDS-PAGE. Maximum GAD activity was observed at pH 5.0 and 55℃ and the activity was dependent on pyridoxal 5'-phosphate. The K_m and V_max of GAD were 0.045 mM and 0.011 mM/min, respectively, when glutamate was used as the substrate.

• Journal of Microbiology and Biotechnology
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• 제19권4호
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• pp.362-367
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• 2009

In recombinant strains, many proteins and enzymes are expressed as inactive and insoluble inclusion bodies. For soluble expression of an active form of StyB, an NADH-flavin oxidoreductase, several recombinant Escherichia coli strains were developed and tested. Among them, strain BL21(DE3)pLysS effectively produced an active and soluble form of StyB as about 9% of the total protein content, when cultivated at $20^{\circ}C$ with 0.5 mM IPTG. The solubly expressed StyB has the highest oxidoreductase activity at pH 6.5-7.5 and $37^{\circ}C$. Substrate dependence profiles of the StyB-catalyzed reaction showed that the maximum specific activity($V_m$) and half saturation constant($K_m$) were $1,867{\pm}148\;U/mg$ protein and $51.6{\pm}11{\mu}M$ for NADH, and $1,274{\pm}34\;U/mg$ protein and $8.2{\pm}1.2{\mu}M$ for FAD, respectively. This indicates that solubly produced StyB has 6- to 9-fold higher oxidoreductase activities than the in vitro refolded StyB from inclusion bodies.