• 한국생물물리학회:학술대회논문집
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• 한국생물물리학회 2003년도 정기총회 및 학술발표회
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• pp.44-44
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• 2003

$BK_{Ca}$ channels were suggested to contain one or more domains of the ‘regulator of K+ conductance’(RCK) in their cytosolic carboxyl termini (Jiang et al.2001). It was also shown that the RCK domain in mammalian $BK_{Ca}$ channels might sense the intracellular $Ca^{2+}$ with a low affinity (Xia et al. 2002). We aligned the amino acid sequence of the $\alpha$-subunit of rat $BK_{Ca}$ channels (rSlo) with known RCK domains and identified a second region exhibiting about 50％ homology. This putative domain, RCK2, contains the characteristic amino acids conserved in other RCK domains. We wondered whether this second domain is involved in the domain-domain interaction and the gating response to intracellular $Ca^{2+}$ for rSlo channel, as revealed in the structure of RCK domain of E. coli channel (Jiang et al.2001). In order to examine the possibility, site-directed mutations were introduced into the RCK2 domain of rSlo channel and the mutant channels were expressed in Xenopus oocytes for functional studies. One of such mutation, G772D, in the putative nucleotide-binding domain resulted in the enhanced $Ca^{2+}$ sensitivity and the channel gating of rSlo channel. These results suggest that this region of $BK_{Ca}$ channels is important for the channel gating and may form an independent domain in the cytosolic region of $BK_{Ca}$ channels. In order to obtain the mechanistic insights of these results, G772 residue was randomly mutagenized by site-directed mutagenesis and total 17 different mutant channels were constructed. We are currently investigating these mutant channels by electrophysiological techniques.ical techniques.

• 식물분류학회지
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• 제38권1호
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• pp.51-61
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• 2008

잎은 무한생장기관으로 잎의 극성제어에 많은 유전적인 요소가 필요하다. 이들 극성은 잎의 초기발생과정에서 제어되기 시작하고, 정단분열조직과 잎기관의 원기와의 제어를 담당하는 인자들에 의해서 결정이 된다. 본 연구에서는 가늘고 바늘처럼 생긴 잎을 가진 deformed root and leaf1 (drl1) 돌연변이체를 유전학적 해석하였고, 그 결과 DRL1 유전자는 정단분열조직과 잎의 극성축을 제어하고 있는 것으로 판명되었다. 이 DRL1 유전자는 효모의 KTI12 유전자 산물과 유사한 단백질인 Elongator associate protein을 만들어 내는 것으로 판명되었다. 또한, 이 단백질의 아미노산 서열이 원핵생물에서부터 진핵생물까지 광범위하게 진화적으로 보존되고 있는 것으로 밝혀졌다. 특히, DRL1 단백질과 유사한 식물의 단백질은 계통해석 결과 단일계통을 나타내고 있는 것으로 나타났고, 이는 이 단백질들이 육상식물의 진화과정에서 잘 보존되고 있음을 시사하고 있다.

• BMB Reports
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• 제44권6호
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• pp.387-392
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• 2011

To investigate the molecular scavenging capabilities of the larvae of Hermetia illucens, two serine proteases (SPs) were cloned and characterized. Multiple sequence alignments and phylogenetic tree analysis of the deduced amino acid sequences of Hi-SP1 and Hi-SP2 were suggested that Hi-SP1 may be a chymotrypsin- and Hi-SP2 may be a trypsin-like protease. Hi-SP1 and Hi-SP2 3-D homology models revealed that a catalytic triad, three disulfide bonds, and a substrate-binding pocket were highly conserved, as would be expected of a SP. E. coli expressed Hi-SP1 and Hi-SP2 showed chymotrypsin or trypsin activities, respectively. Hi-SP2 mRNAs were consistently expressed during larval development. In contrast, the expression of Hi-SP1 mRNA fluctuated between feeding and molting stages and disappeared at the pupal stages. These expression pattern differences suggest that Hi-SP1 may be a larval specific chymotrypsin-like protease involved with food digestion, while Hi-SP2 may be a trypsin-like protease with diverse functions at different stages.

• Journal of Ginseng Research
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• 제44권2호
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• pp.321-331
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• 2020

Background: The patatin-related phospholipase AIII family (pPLAIIIs) genes alter cell elongation and cell wall composition in Arabidopsis and rice plant, suggesting diverse commercial purposes of the economically important medicinal ginseng plant. Herein, we show the functional characterization of a ginseng pPLAIII gene for the first time and discuss its potential applications. Methods: pPLAIIIs were identified from ginseng expressed sequence tag clones and further confirmed by search against ginseng database and polymerase chain reaction. A clone showing the highest homology with pPLAIIIβ was shown to be overexpressed in Arabidopsis using Agrobacterium. Quantitative polymerase chain reaction was performed to analyze ginseng pPLAIIIβ expression. Phenotypes were observed using a low-vacuum scanning electron microscope. Lignin was stained using phloroglucinol and quantified using acetyl bromide. Results: The PgpPLAIIIβ transcripts were observed in all organs of 2-year-old ginseng. Overexpression of ginseng pPLAIIIβ (PgpPLAIIIβ-OE) in Arabidopsis resulted in small and stunted plants. It shortened the trichomes and decreased trichome number, indicating defects in cell polarity. Furthermore, OE lines exhibited enlarged seeds with less number per silique. The YUCCA9 gene was downregulated in the OE lines, which is reported to be associated with lignification. Accordingly, lignin was stained less in the OE lines, and the expression of two transcription factors related to lignin biosynthesis was also decreased significantly. Conclusion: Overexpression of pPLAIIIβ retarded cell elongation in all the tested organs except seeds, which were longer and thicker than those of the controls. Shorter root length is related to auxinresponsive genes, and its stunted phenotype showed decreased lignin content.

• Journal of Microbiology
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• 제42권2호
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• pp.152-155
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• 2004

Pseudomonas sp. K82 has been reported to be an aniline-assimilating soil bacterium. However, this strain can use not only aniline as a sole carbon and energy source, but can also utilize benzoate, p-hydroxybenzoate, and aniline analogues. The strain accomplishes this metabolic diversity by using dif-ferent aerobic pathways. Pseudomonas sp. K82, when cultured in p-hydroxybenzoate, showed extradiol cleavage activity of protocatechuate. In accordance with those findings, our study attempted the puri-fication of protocatechuate 4,5-dioxygenase (PCD 4,5). However the purified PCD 4,5 was found to be very unstable during purification. After Q-sepharose chromatography was performed, the crude enzyme activity was augmented by a factor of approximately 4.7. From the Q-sepharose fraction which exhibited PCD 4,5 activity, two subunits of PCD4,5 (${\alpha}$ subunit and ${\beta}$ subunit) were identified using the N-terminal amino acid sequences of 15 amino acid residues. These subunits were found to have more than 90％ sequence homology with PmdA and PmdB of Comamonas testosteroni. The molecular weight of the native enzyme was estimated to be approximately 54 kDa, suggesting that PCD4,5 exists as a het-erodimer (${\alpha}$$_1$${\beta}$$_1$). PCD 4,5 exhibits stringent substrate specificity for protocatechuate and its optimal activity occurs at pH 9 and 15 $^{\circ}C$. PCR amplification of these two subunits of PCD4,5 revealed that the ${\alpha}$ subunit and ${\beta}$ subunit occurred in tandem. Our results suggest that Pseudomonas sp. K82 induced PCD 4,5 for the purpose of p-hydroxybenzoate degradation.

• 한국미생물·생명공학회지
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• 제42권2호
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• pp.184-189
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• 2014

본 연구에서는 강원전통장류로부터 혈전용해 활성이 우수한 효모를 분리하였다. 된장에서 분리한 AFY-1 균주는 혈전용해 활성 측정결과 양성대조구인 plasmin 보다 약 1.75배 높은 활성을 나타내었다. 분리한 효모는 18S rRNA 염기서열 및 탄소원 이용 특성 분석을 통하여 Saccharomycetales sp.로 동정되었으며, AFY-1 균주의 생육 최적온도는 $32^{\circ}C$였다. 본 연구에서 분리된 혈전용해능이 우수한 효모균주는 안전성 검증 등 추가연구를 통해 발효식품 제조용 스타터로서 활용이 가능할 것으로 기대된다.

• 한국정보과학회논문지:소프트웨어및응용
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• 제31권12호
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• pp.1602-1610
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• 2004

다양한 유전체 프로젝트로 말미암아 엄청난 서열 데이타들이 쏟아지고, 이에 따라 핵산 및 단백질 수준의 서열 데이타 분석이 매우 중요하게 인식되고 있다. 특히 최근에는 단백질이 단순하게 개별적인 기능을 가진 독립적인 요소가 아닌 전체 단백질 상호작용 네트워크 상에서 다른 객체들과 유기적인 관계를 갖으며 나름대로의 중요한 역할을 수행하고 있다는 점에 초점을 맞추어 연구가 진행되고 있다. 특히 단백질 상호작용 관계 분석을 위해서는 실제로 상호작용이 일어나는 도메인 모듈 정보가 아주 중요하게 작용하는데, 본 논문에서는 이러한 점을 고려하여 우리가 개발한 단백질 기능 및 상호작용 분석을 위한 PIVS(Protein-protein interaction Inference and Visualization System)에 대해 소개하고 있다 PIVS는 기존의 단백질 상호작용 데이타베이스들을 합쳐서 통합 데이타베이스를 생성하고, 다양한 전처리 과정으로 통합 데이타베이스 서열의 기능 및 주석 정보를 추출하여 로컬 데이타베이스화 하였다. 그리고 특히 단백질 상호작용 관계 분석을 위해 중요하게 작용하는 도메인 모듈 정보들을 추출하여 로컬 데이터베이스를 구축하였고, 기존의 단백질 상호작용 관계 데이타를 이용하석 도메인 사이의 상호작용 관계 정보도 수집하여 분석하였다. PIVS는 단백질의 종합적인 분석 정보, 즉, 기능 및 주석, 도메인, 상호작용 관계 정보 등을 손쉽고 편리하게 얻고자 하는 사용자에게 매우 유용하게 사용될 수 있을 것이다.

• Applied Biological Chemistry
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• 제40권5호
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• pp.388-394
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• 1997

곤충 병원성 곰팡이 Beauveria bassiana ATC7159의 배양여액으로부터 균체외 protease (basiasin I)를 Ammonium sulfate 침전, DEAE-Sephadex A-50, CM-cellulose 및 Hydroxyapatite column chromatography를 수행하여 완전히 정제하였다. 이 과정으로 41배 정제되었으며 정제수율은 13.6%였다. 정제된 bassiasin I의 분자량은 SDS-PAGE 상에서 약 32,000 Da이며 pI값은 9.5로 확인되었다. $NH_2$ 말단 아미노산 서열은 다른 곤충 병원성 곰팡이가 생산하는 pretense들과 높은 상동성을 보였다. Protease 활성의 최적 pH는 10.5 부근이며, pH 5.0-11.0범위에서 안정하였다. 최대 활성온도는 $60-65^{\circ}C$이며, $60^{\circ}C$에서 120분후에 약 20%의 잔존활성을 보였다. 이 pretense는 phenylmethylsulfonyl fluoride (PMSF) 및 diisopropyl fluorophosphate (DFP)에 의해 저해된다.

• Mycobiology
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• 제48권5호
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• pp.427-430
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• 2020

Ganoderma lingzhi is a well-known source of natural fungal medicines which has been given for the treatment of several diseases. China is one of the major commercial producers of Ganoderma mushroom worldwide. However, with the expansion of the commercial cultivation, the occurrence of the fungal diseases on G. lingzhi has also been increased. The green mold disease symptoms were observed in the cultivation base of G. lingzhi in Zuojia Town, Jilin City, Jilin Province, China, causing the basidiomes to be rotten and withered, and the green mycelium layer generated gradually. The pathogenicity tests showed the same symptoms as appeared naturally in Zuojia mushroom base. Morphology characters revealed conidia green, ellipsoid, globose, 2.56-4.83 × 2.09-4.22 ㎛, length-width ratio was 1.1-1.2 (n = 10). Conidiophores trichoderma-like, often asymmetry, branches solitary, paired or in whorls of 3 phialides formed solitary, paired or in whorl, variable in shape, lageniform, sometimes ampulliform or subulate. While using molecular methodology, comparing with the sequences of Trichoderma hengshanicum from GenBank, the analyzed sequence showed 97.32% homology with the RPB2 sequences, 100% with the TEF1-a sequences. A fungus isolated from the diseased tissues was identified based on morphology and molecular studies as T. hengshanicum. This is the first report of T. hengshanicum causing the green mold disease of G. lingzhi in China.

• Biomolecules & Therapeutics
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• 제25권2호
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• pp.171-176
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• 2017

Streptomyces avermitilis produces clinically useful drugs such as avermectins and oligomycins. Its genome contains approximately 33 cytochrome P450 genes and they seem to play important roles in the biosynthesis of many secondary metabolites. The SAV_7130 gene from S. avermitilis encodes CYP158A3. The amino acid sequence of this enzyme has high similarity with that of CYP158A2, a biflaviolin synthase from S. coelicolor A3(2). Recombinant S. avermitilis CYP158A3 was heterologously expressed and purified. It exhibited the typical P450 Soret peak at 447 nm in the reduced CO-bound form. Type I binding spectral changes were observed when CYP158A3 was titrated with myristic acid; however, no oxidative product was formed. An analog of flaviolin, 2-hydroxynaphthoquinone (2-OH NQ) displayed similar type I binding upon titration with purified CYP158A3. It underwent an enzymatic reaction forming dimerized product. A homology model of CYP158A3 was superimposed with the structure of CYP158A2, and the majority of structural elements aligned. These results suggest that CYP158A3 might be an orthologue of biflaviolin synthase, catalyzing C-C coupling reactions during pigment biosynthesis in S. avermitilis.