Effect of Interaction between Protocatechualdehyde Produced from Streptomyces lincolnensis M-20 and Copper Ions on Antioxidant and Pro-oxidant Activities

김경자,이재훈,양용준,Kim, Kyoung-Ja,Lee, Jae-Hun,Yang, Yong-Joon The Microbiological Society of Korea 2014 미생물학회지 Vol.50 No.1

Protocatechualdehyde (PA)는 항산화 활성과 항암 활성을 가진 페놀성 물질이다. Streptomyces lincolnensis M-20 균주에서 생산된 PA를 균주 상등액에서 분리, 정제하였다. 항산화 활성을 가진 PA가 구리 이온 존재 하에서는 산화촉진제로 작용하였다. 항산화 활성은 DPPH를 이용한 방법으로 측정하였으며, 구리 이온 존재 하에서 PA의 산화 촉진 작용은 pBR322 플라스미드의 DNA 절단 작용으로 측정하였다. DNA 손상으로 생성되는 활성산소 종의 확인은 활성 산소종의 포집자인 글루타치온에 의해 DNA 절단이 억제되는 것으로 확인하였다. PA와 구리 이온의 복합체 형성은 금속 이온의 킬레이트인 EDTA가 존재할 경우와 존재하지 않을 경우를 자외선/가시광선 분광학적 분석법으로 비교, 확인하였다. Protocatechualdehyde (PA) is phenolic compound having antioxidative and antitumor activities. PA was purified from supernatant of Streptomyces lincolnensis M-20. In the presence of copper ion, PA acted as pro-oxidant. The antioxidant activity was assessed with the 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay, and the pro-oxidant effect of PA on DNA damage as pBR322 plasmid DNA-cleaving agents in the presence of Cu(II) ions was investigated. The involvement of reactive oxygen species (ROS) in the DNA damage was confirmed by the inhibition of the DNA breakage by using glutathione (GSH), specific scavenger of ROS. When the increase in ROS reaches a certain level (the toxic threshold), it may trigger cell death. The formation of the PA/Cu(II) chelate complex was confirmed by reaction with ethylenediamine-tetraacetic acid (EDTA), a well-known chelating agent for metal ions, by using UV/Vis spectroscopic analysis.

Production of siderophore from L-glutamic acid as both carbon and nitrogen sole sources in Acinetobacter sp. B-W

김경자,장주호,양용준,Kim, Kyoung-Ja,Jang, Ju-Ho,Yang, Yong-Joon The Microbiological Society of Korea 2017 미생물학회지 Vol.53 No.2

포도당과 글루탐산을 함유한 배지에서 시드로포어인 2, 3-dihydroxybenzoic acid (DHB)를 생산하는 Acinetobacter sp. B-W 균주를 글루탐산을 유일한 탄소원과 질소원으로 함유한 배지에 배양한 결과, 상등액에서 2, 3-DHB가 아닌 카테콜 형의 시드로포어를 생산하는 것으로 조사되었다. 글루탐산의 농도는 3%에서 시드로포어 생산이 최고로 나타났으며, 3% 보다 높은 농도에서는 감소하는 것으로 조사되었다. 글루탐산을 유일한 탄소원과 질소원으로 함유한 배지에서 자란 균주 B-W는 배양 온도 $28^{\circ}C$에서는 시드로포어를 생산하지만 $36^{\circ}C$에서는 생산하지 않았다. 또한 $10{\mu}M\;FeCl_3$를첨가한 배지에서는 시드로포어 생산이 완전히 억제되었다. 글루탐산 배지에서 생산된 균주 B-W의 시드로포어는 TLC 전개 용매 butanol: acetic acid: water =12:3:5에서 Rf치가 0.32로 나타나 Rf치가 0.82인 2, 3-DHB와는 다른 것으로 조사되었으며, 또한 항산화 활성도 없는 것으로 나타나 항산화 활성을 가진 2, 3-DHB와는 다른 시드로포어인 것으로 추정되었다. Catechol type siderophore different from 2, 3-dihydroxybenzoic acid (DHB) was produced from Acinetobacter sp. B-W grown in medium containing L-glutamic acid as both carbon and nitrogen sole sources at $28^{\circ}C$. Optimal concentration of glutamic acid for siderophore production was 3% and production of siderophore was decreased above 3% glutamic acid. In previous report, siderophore, 2, 3-DHB was produced from strain B-W grown in medium containing glucose as carbon source and glutamic acid as nitrogen source. Rf value of siderophore produced from strain B-W grown in medium glutamic acid as both carbon and nitrogen sole sources at $28^{\circ}C$ was 0.32, while 2, 3-DHB was 0.84 in butanol-acetic acid-water (12:3:5) as developing solvent. Antioxidative activity of 2, 3-DHB was not detected in that siderophore produced from glutamic acid. Catechol nature of siderophore was detected by Arnow test. Although in iron-limited media optimal cell growth was identified at $36^{\circ}C$, significant quantities of siderophore were produced only at $28^{\circ}C$. Biosynthesis of siderophore was strongly inhibited by growth at $36^{\circ}C$. Production of siderophore was completely inhibited by $10{\mu}M\;FeCl_3$.

Effect of plasmid curing on the production of siderophore from glutamic acid as both carbon and nitrogen sole sources in Acinetobacter sp. B-W

김경자,이재림,양용준,Kim, Kyoung-Ja,Lee, Jae-Rim,Yang, Yong-Joon The Microbiological Society of Korea 2018 미생물학회지 Vol.54 No.3

Effect of plasmid curing of Acinetobacter sp. B-W on the production of siderophore from glutamic acid as both carbon and nitrogen sole sources was investigated. Plasmid cured mutant of strain B-W lost the ability to produce siderophore from glutamic acid at $28^{\circ}C$. Transformant E. coli $DH5{\alpha}$ harboring 20 kb plasmid, that was isolated from wild type of strain B-W produced siderophore from glutamic acid as both carbon and nitrogen sole sources at $28^{\circ}C$, but, not at $36^{\circ}C$. Production of siderophore from glutamic acid by transformant E. coli $DH5{\alpha}$ was completely inhibited by $10{\mu}M\;FeCl_3$. In previous report, catechol nature of siderophore produced from glutamic acid by strain B-W was detected by Arnow test. The siderophore produced from glutamic acid by transformant E. coli $DH5{\alpha}$ was also catechol type. Rf value of siderophore produced from transformant E. coli $DH5{\alpha}$ grown in medium glutamic acid as both carbon and nitrogen sole sources at $28^{\circ}C$ was 0.32 in butanol-acetic acid-water (12:3:5) as developing solvent. Rf value of the siderophore was the same with that of wild type of strain B-W. Thus a single plasmid of 20 kb seemed to be involved in the production of siderophore from glutamic acid. 플라스미드가 제거된 Acinetobacter sp. B-W 균주의 돌연변이체를 글루탐산을 유일한 탄소 원과 질소원으로 함유한 배지에 $28^{\circ}C$에서 배양한 결과 글루탐산으로부터의 시드로포어 생산이 억제되었다. B-W 원 균주의 20 kb 플라스미드를 가진 형질 전환체 대장균 $DH5{\alpha}$는 같은 조건의 배지에서 시드로포어를 생산하는 것으로 조사되었다. 그러나 $36^{\circ}C$에서는 형질 전환체 대장균 $DH5{\alpha}$의 시드로포어 생산이 강하게 억제되었으며, 돌연변이체 B-W 균주는 $28^{\circ}C$에서와 마찬가지로 $36^{\circ}C$에서도 시드로포어를 생산하지 못하였다. 형질 전환체로부터 생산된 시드로포어의 종류는 원 균주 B-W와 마찬가지로 Arnow 시험 결과 카테콜 형으로 조사되었으며, $10{\mu}M\;FeCl_3$를 첨가한 배지에서는 시드로포어 생산이 완전히 억제되었다. 형질전환체로부터 생산된 시드로포어의 TLC상에서의 Rf값은 butanol-acetic acid-water (12:3:5) 용매상에서 0.32로 원 균주 B-W에서 생산된 시드로포어와 같았다. 위와 같은 실험 결과로 글루탐산으로부터 생산된 시드로포어의 생합성에 관여하는 유전자들이 20 kb 플라스미드 상에 있는 것으로 추정되었다.

Strepfomyces lincolnensis M-20 균주로 부터 분리, 정제된 L-Asparaginase의 열안정성과 단백 가수 분해 효소에 대한 저항성

김경자,Kim, Kyoung-Ja 대한약학회 2007 약학회지 Vol.51 No.3

Thermostable asparaginase was purified to homogeneity from mesophilic Strepfomyces lincolnensis M-20 by 30${\sim}$70% ammonium sulfate precipitation and asparagine-Sepharose CL 6B affinity column chromatography, The apparent molecular mass of L-asparaginase by SDS-PAGE was found to be 47 kDa, whereas by its mobility on Sephacryl S-300 column was around 180 kDa, indicating that the enzyme at the native stage acts as tetramer, The purified enzyme showed a single band on acrylamide gel electrophoresis. The optimum pH and temperature were pH 9.5 and 55${\circ}$C, respectively. Chemical modification experiments of purified asparagines implied the existence cystein residue located at or near active site. Purified asparaginase retained the 85% of the initial activity after incubation at 90${\circ}$C for 30 min. A correlation between themostability and resistance to proteolysis of commercial asparaginase and purified asparaginase from Strepfomyces lincolnensis M-20 was investigated. Purified thermostable asparaginase was resistant to trypsin and chymotrypsin treatment, while the commercial asparaginase was not themostable and was susceptible to proteolytic treatment with trypsin and chymotrypsin.

가용 시간을 이용한 역할 기반 위임 모델

김경자,장태무,Kim, Kyoung-Ja,Chang, Tae-Mu 한국정보처리학회 2007 정보처리학회논문지 C : 정보통신,정보보안 Vol.14 No.1

기존의 RBAC(Role Based Access Control) 모델들은 사용자간의 역할 위임이나 역할 분리를 운용하기에는 역부족이다. 이에 RBAC을 바탕으로 역할이나 권한을 다른 사용자에게 위임하는 모델인 RBDM(Roie Based Delegation Model)이 연구되고 있다. 반면에, 역할을 위임 받은자의 악의적인 남용을 막을 수 있는 방안이 필요하다. 본 논문은 역할의 위임으로 인하여 위임 받은자의 악의적인 도용을 막기 위한 모델로서 위임 역할에 대한 폐지 절차를 시간상으로 제한을 두고, 위임하는 역할에 대하여 원사용자의 권한을 그대로 유지함으로써 위임으로 인한 역할간 계층구조의 보안상 취약점을 보완하고자 한다. 즉, 역할을 위임하고자 할때에는 위임에 대한 유효 기간을 같이 부여하는 기법으로 위임된 역할의 사용 권한을 시간상으로 폐지할 수 있게 한다. 또한, 위임된 역할에 대해서는 원사용자의 역할에 대한 권한을 계속 보유할 수 있도록 함으로써 위임으로 인하여 역할에 대한 사용이 정당하지 않은 경우에는 언제든지 회수가 가능할 수 있게 하였다. 본 제안 기법인 T-RBDM (Time-out based RBDM)은 기존의 위임 모델인 RBDM0, RBDM1, PBDM과 비교 분석함으로 기존의 위임 모델에 비해 보안 측면에서 더 강인함을 보인다. The existing RBAC models are not sufficient for managing delegations or separation of roles. Researches have been done on RBDM(Role Based Delegation Model) that deal with delegating role or permission to other users. In this paper, we divide the delegated roles into two groups: periodic and temporary delegation roles. When a role is delegated, a time period is assigned together, which is used to revoke the permission of delegated role automatically. In our model, the role of monotonic delegation by an original user can be revoked at any time in case of malicious use by the delegated user. The contribution of our model is that the malicious use of delegated role can be prohibited and security vulnerability in the role hierarchy due to role delegations can be alleviated. The proposed model, T RBDM(Time out Based RBDM) is analyzed and compared with the conventional models, such as RBDM0, RBDM1 and PBDM. Our model shows an advantage over other models in terms of security robustness.

Temperature dependent 2,3-dihydroxybenzoic acid production in Acinetobacter sp. B-W

김경자,이재훈,양용준,Kim, Kyoung-Ja,Lee, Jae-Hun,Yang, Yong-Joon The Microbiological Society of Korea 2015 미생물학회지 Vol.51 No.3

A soil microorganism producing iron chelator (siderophore) under low iron stress (up to $2{\mu}M$ of iron) was identified as Acinetobacter sp. B-W by 16S rDNA sequence analysis, biochemical-, physiological tests and morphological analysis using electron microscope. Catechol nature of siderophore was detected by Arnow test. Although optimal cell growth was identified at $36^{\circ}C$ in iron-limited media, significant quantities of siderophore were produced only at $28^{\circ}C$. Biosynthesis of siderophore was strongly inhibited by growth at $36^{\circ}C$. Production of siderophore was completely inhibited by $10{\mu}M\;FeCl_3$. Iron chelator produced from Acinetobacter sp. B-W was purified from supernatant using butanol extraction, Sephadex LH-20 column chromatography and HPLC. Purified sideropore was identified as 2,3-dihydroxybenzoic acid by HPLC, TLC and IR analysis. 철 스트레스($2{\mu}M$ 이하 농도) 하에서 시데로포어를 생산하는 균주를 토양에서 분리하여 16S rDNA 염기 서열 분석과 생화학적, 생리학적 분석 및 전자 현미경 관찰 등으로 동정한 결과, Acinetobacter sp.로 밝혀졌다. 시데로포어의 카테콜 특성은 Arnow법으로 조사되었다. 철을 제한한 배지에서 균주를 배양한 결과, $36^{\circ}C$에서도 잘 자랐지만 시데로포어 생산은 $28^{\circ}C$에서 높았다. $36^{\circ}C$에서는 시데로포어 생산이 강하게 억제되었다. $10{\mu}M\;FeCl_3$를 첨가한 배지에서는 시데로포어 생산이 완전히 억제되었다. 균주 상등액을 부탄올 추출 후, Sephadex LH-20 컬럼 크로마토그래피와 HPLC를 이용하여 시데로포어를 분리, 정제하였다. 분리, 정제된 시데로포어의 구조는 HPLC, TLC와 IR 분석 결과로부터 2,3-dihydroxybenzoic acid로 확인되었다.

Burkholderia sp. CAS-5 균으로 부터 생산된 시드로포어의 Matrix metalloproteinase-2(Gelatinase A) 억제 활성

김경자,Kim, Kyoung-Ja 대한약학회 2006 약학회지 Vol.50 No.4

Matrix metalloproteinase-2 is known to be involved in pathological processes such as tumor invasion or rheumatoid arthritis. A soil microorganism producing siderophore under low iron stress $(up\;to\;5\;{\mu}m\;of\;iron)$ was identified as Burkholderia sp. Hydroxamate type siderophore produced by Burkholderia sp. CAS-5 was partially purified. MMP inhibitory activity of siderophore was confirmed by gelatin zymography. The $Zn^{2+}-chelating$ activity of siderophore correlated with the inhibition of MMP-2 activity.

애드혹 네트워크에서 ZRP를 기반으로 하는 경로 탐색 기법

김경자,장태무,Kim, Kyoung-Ja,Chang, Tae-Mu 한국정보처리학회 2004 정보처리학회논문지 C : 정보통신,정보보안 Vol.11 No.3

애드혹 네트워크는 고정된 기반 구조 없이 이동 호스트들로만 구성된 네트워크이다. 이러한 네트워크는 노드의 이동으로 인한 토폴로지의 잦은 변화로 관리면에서 많은 어려움이 있다. 따라서, 애드혹 네트워크의 라우팅 경로(Routing Path)에 대한 신뢰도를 높이는 것이 점점 중요해 지고 있다. 본 논문에서는 기존의 ZRP(Zone Routing Protocol)를 기반으로 하여 라우팅 경로의 전체적인 홉(hop) 수를 줄이고, 경로상의 노드들끼리의 인증을 통하여 라우팅 경로의 신뢰성을 향상시키는 경로 탐색 방안을 제안하였다. 또한 토폴로지의 잦은 변화로 인한 라우팅 경로 유지의 어려움을 질의 제어 메커니즘을 통하여 해결하고자 시도하였다. 본 논문의 방안은 시뮬레이션을 통하여 그 효율성을 입증할 수 있었다. Ad hoc networks are groups of mobile hosts without any fixed infrastructure. Frequent changes in network topology owing to node mobility make these networks very difficult to manage. Therefore, enhancing the reliability of routing paths in ad hoc networks gets more important. In this paper, we propose a ZRP(Zone Routing Protocol)-based route discovery scheme that can not only reduce the total hops of routing path, but Improve security through authentications between two nodes. And to solve the problem in maintenance of routing paths owing to frequent changes of the network topology, we adopt a query control mechanism. The effectiveness of our scheme is shown by simulation methods.

장기요양시설 요양보호사의 감염관리 수행에 영향을 미치는 요인

김경자,박성원,Kim, Kyoung Ja,Park, Sung Won 한국가정간호학회 2016 가정간호학회지 Vol.23 No.2

Purpose: The purpose of this study was to investigate factors influencing health workers' infection management behavior in long-term care facilities. Methods: A descriptive cross-sectional survey was conducted with 180 health workers who are employed in long-term care facilities. The data were collected from April, 25 until July, in 2016. Results: Infection management behavior positively correlated with the perceived importance of infection management (r=.77, p<.001), but role conflict negatively correlated with infection management behavior (r=.28, p<.001). The hierarchical regression model with general characteristics (first step) and perceived importance of infection management, work environment, and role conflict (second step) against infection management behavior was statistically significant (F=31.93, p<.001). This model could explain 62.8% of infection management behavior ($R^2=.62$, ${\Delta}R^2=.39$). Particularly, perceived importance of infection management was identified as factors influencing infection management behavior(${\beta}=.70$, p<.001). Conclusion: The findings of this study imply that systemic education about infection control and monitoring should be considered, so as to encourage proper infection management behaviors among health workers in long-term care facilities.

Streptomyces sp. M-20 균의 대사물에 의한 Streptococcus mutans의 Glucosyltransferase 활성 억제 효과

김경자,Kim, Kyoung-Ja 대한약학회 2005 약학회지 Vol.49 No.2

Dental caries is one of the most common oral diseases in the world. Glucosyltransferase (Gtase) of Streptococcus mutans (S. mutans) plays an important role in the develo pment of dental caries. For the purpose to develop anti- caries, we examined the effect of metabolites isolated from Streptomyces sp. M-20 on Gtase and the growth of S. mutans. Streptomyces sp. M-20 isolated from Mongolian soil showed 95~96% sequence homology with that of Streptomyces lin- colnensis. The metabolites of Streptomyces sp. M-20 were partially purified by extraction with ethyl acetate, silica gel column chromatography and preparative TLC. Partially purified metabolite, red colored component (MR-20) in ethyl acetate fraction showed potent antibacterial activitiy against S. mutans and inhibitory activity against Gtase purified from S. mutans, while another isolated yellow component (MY-20) showed no activity against S. mutans. The inhibitory activity of MR-20 against Gtase was confirmed by activity staining on SDS-polyacrylamide gel electrophoresis. The concentration of MR-20 for 50% inhibition $(IC_{50})$ against Gtase activity was $60{\mu}g/ml$. These results suggest that MR-20 can be developed for antibacterial agent and anticaries.