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Glucose Oxidase의 Sulfhydryl Group 특성 연구 : 5,5'-dithiobis(2-nitrobenzoate)와의 반응성
김태련,고은희,Kim, Tae-Ryun,Koh, Eun-Hie 생화학분자생물학회 1986 한국생화학회지 Vol.19 No.4
Glucose oxidase의 sulfhydryl group의 분자내 특성을 추정하기 위하여 5,5'-dithiobis(2-nitrobenzoate) (DTNB)와의 반응을 연구하였다. DTNB는 유리된 -SH와 반응하여 412 nm에서 정량할 수 있으며 이와 동시에 반응속도를 측정하므로써 분자내에서의 -SH group의 환경을 추정할 수 있다. Glucose oxidase의 활동도는 lactoperoxidase와 O-dianisidine을 사용한 coupled enzyme system을 이용하였다. 천연효소(native form)는 DTNB와 매우 느리게 반응하여 (표준조건에서 $5.6{\times}10^{-5}\;s^{-1}$) -SH group이 쉽게 곁에서 접근할 수 없음을 보여주고 있다. 각종 denaturing 시약으로 효소를 비활성화 시킴에 따라 DTNB와의 반응속도가 그 시약의 특성에 따라 증가함을 관찰할 수 있었다. 4 M urea에서는 1.9개의 -SH group이 정량되었고 속도는 $1.7{\times}10^{-3}\;s^{-1}$이었다. 한편 좀 더 미세한 분자내 환경을 관찰하기 위하여 여러 종류의 disulfide를 매체로 한 DTNB와의 반응속도를 측정했으며 그 결과 cystamine은 DTNB와의 반응을 촉진시키는 것으로 나타났다. The molecular characteristics of sulfhydryl groups of glucose oxidase was studied by the reaction of DTNB. DTNB reacts with free -SH groups of proteins to generate a chromophore which can be measured spectrophotometrically and the DTNB reaction rates can be used to estimate the microenvironment of the -SH group in the molecule. The enzymatic activity of glucose oxidase was determined by measuring the coupled enzyme reaction with lactoperoxidase and O-dianisidine. It was observed that the reaction of glucose oxidase with DTNB was very slow (kobs was $5.6{\times}10^{-5}\;s^{-1}$ under the standard condition) when the enzyme was in the native form. This indicates that the -SH group are not easily approachable from the surface. When various denaturing agents were added, the DTNB reaction rate increased according to the specific properties of added denaturing agents. The DTNB reaction rate in the presence of 4 M urea was $1.7{\times}10^{-3}\;s^{-1}$ and there are 1.9 titrable sulfhydryl groups. Additionally, the DTNB reactions of glucose oxidase mediated by various disulfides were studied in order to observe the minute microscopic molecular environment. Among those studied, cystamine seems to increase the DTNB reaction noticeably.
김태련,양철학,Kim, Tae Ryeon,Yang, Cheol Hak Korean Chemical Society 1995 Bulletin of the Korean Chemical Society Vol.16 No.5
The P. fragi lipase overexpressed in E. coli as a fusion protein of 57 kilodalton (kDa) has been purified through glutathione-agarose affinity chromatography by elution with free glutathione. The general properties of the purified GST-fusion protein were characterized by observing absorbance of released p-nitrophenoxide at 400 nm which was hydrolyzed from the substrate p-nitrophenyl palmitate. The optimum condition was observed at 25 $^{\circ}C$, pH 7.8 with 0.4 ${\mu}g$ of protein and 1.0 mM substrate in 0.6% (v/v) TritonX-100 solution. Also the lipase was activated by Ca+2, Mg+2, Ba+2 and Na+ but it was inhibited by Co+2 and Ni+2. pGEX-2T containing P. fragi lipase gene as expression vector was named pGL191 and used as a template for the site-directed mutagenesis by sequential PCR steps. A Ser-His-Asp catalytic triad similar to that present in serine proteases may be present in Pseudomonas lipase. Therefore, the PCR fragments replacing Asp217 to Arg and His260 to Arg were synthesized, and substituted for original fragment in pGL19. The ligated products were transformed into E. coli NM522, and pGEX-2T harboring mutant lipase genes were screened through digestion with XbaI and StuI sites created by mutagenic primers, respectively. No activity of mutant lipases was observed on the plate containing tributyrin. The purified mutant lipases were not activated on the substrate and affected at pH variation. These results demonstrate that Asp217 and His260 are involved in the catalytic site of Pseudomonas lipase.
오행침자법(五行鍼刺法)을 포함한 한방치료로 호전된 뇌경색 후 발생한 Broca 실어증 치험 1례
김태련,임현찬,전상윤,Kim, Tae-ryun,Lim, Hyun-chan,Jeon, Sang-yun 대한한방내과학회 2016 大韓韓方內科學會誌 Vol.37 No.3
Objective: To report the case of a patient suffering from Broca’s aphasia after cerebral infarction, which was improved by Korean medical treatments, especially five-element acupuncture.Methods: To relievesymptoms, Korean medicine treatments such as acupuncture, moxibustion, and herb medication were performed. To evaluate the effect of the treatments, articulation accuracy, vowel accuracy, and the sequential motion rate of articulator were checked.Result: After treatments for 28 days, the symptoms were improved.Conclusion: This report suggests that five-element acupuncture can be effectively used for patients with Broca’s aphasia after cerebral infarction.
Glucose Oxidase 의 Sulfhydryl Group 특성연구 : 5 , 5 ' - dithiobis ( 2 - nitrobenzoate ) 와의 반응성
김태련,고은희 ( Tae Ryun Kim,Eun His Koh ) 생화학분자생물학회 1986 BMB Reports Vol.19 No.4
The molecular characteristics of sulfhydryl groups of glucose oxidase was studied by the reaction of DTNB. DTNB reacts with free -SH groups of proteins to generate a chromophore which can be measured spectrophotometrically and the DTNB reaction rates can be used to estimate the microenvironment of the -SH group in the molecule. The enzymatic activity of glucose oxidase was determined by measuring the coupled enzyme reaction with lactoperoxidase and O-dianisidine. It was observed that the reaction of glucose oxidase with DTNB was very slow (k_(obs) was 5.6×10^(-5)s^(-1) under the standard condition) when the enzyme was in the native form. This indicates that the -SH group are not easily approachable from the surface. When various denaturing agents were added, the DTNB reaction rate increased according to the specific properties of added denaturing agents. The DTNB reaction rate in the presence of 4 M urea was 1.7×10^(-3)s^(-1) and there are 1.9 titrable sulfhydryl groups. Additionally, the DTNB reactions of glucose oxidase mediated by various disulfides were studied in order to observe the minute microscopic molecular environment. Among those studied, cystamine seems to increase the DTNB reaction noticeably.
김태련(Tae Lyun Kim),방희정(Hee Jung Bang),최경숙(Kyoung Sook Choi),조혜자(Hye Ja Cho) 한국아동학회 1991 兒童學會誌 Vol.12 No.2
The present study investigated how the inconsistency between episodes in a story affects judgment of the importance of a sentence. In experiment I, 120 college students read 4 stories containing 3 episodes and judged each of the sentences, one after another. The results of the importance rating showed that adult subjects were sensitive to inconsistency between episodes. The analysis of episodes revealed a significant difference between episodes in all of the 4 stories, with lowest importance ratings given to the inconsistent episodes. In experiment II, 220 children were sampled from a private school of middle S.E.S.Status. 2nd-, 4th- and 6th- graders read 2 stories, and judged the importance of the sentences. It was found that children were less sensitive to inconsistencies in a story than adults. Younger children had more difficulty in the integration of inconsistent information. Especially in the 2nd episode, the importance rating was affected by grade [F(2,2627)=195.52, p<.01], suggesting that 6th graders were more sensitive to inconsistencies than the younger children. In contrast to adults, 2nd graders gave high importance ratings to inconsistent sentences as well as to consistent sentences.