• Journal of Microbiology and Biotechnology
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• 제17권10호
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• pp.1607-1615
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• 2007

We investigated the applicability of the TEM-l ${\beta}$-lactamase fragment complementation (BFC) system to develop a strategy for the screening of protein-protein interactions in bacteria. A BFC system containing a human Fas-associated death domain (hFADD) and human Fas death domain (hFasDD) was generated. The hFADD-hFasDD interaction was verified by cell survivability in ampicillin-containing medium and the colorimetric change of nitrocefin. It was also confirmed by His pull-down assay using cell lysates obtained in selection steps. A coiled-coil helix coiled-coil domain-containing protein 5 (CHCH5) was identified as an interacting protein of human uracil DNA glycosylase (hUNG) from the bacterial BFC cDNA library strategy. The interaction between hUNG and CHCH5 was further confirmed with immunoprecipitation using a mammalian expression system. CHCH5 enhanced the DNA glycosylase activity of hUNG to remove uracil from DNA duplexes containing a U/G mismatch pair. These results suggest that the bacterial BFC cDNA library strategy can be effectively used to identify interacting protein pairs.

• Asian Pacific Journal of Cancer Prevention
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• 제14권11호
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• pp.6803-6806
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• 2013

Background: Interferon regulatory factor 6 (IRF6) is a transcription factor with distinct and conserved DNA and protein binding domains. Mutations within the protein binding domain have been significantly observed in subjects with orofacial cleft relative to healthy controls. In addition, recent studies have identified loss of expression of IRF6 due to promoter hypermethylation in cutaneous squamous cell carcinomas. Since mutational events occurring within the conserved domains are likely to affect the function of a protein, we investigated whether regions within the IRF6 gene that encodes for the conserved protein binding domain carried mutations in oral squamous cell carcinoma (OSCC). Materials and Methods: Total chromosomal DNA extracted from 32 post surgical OSCC tissue samples were amplified using intronic primers flanking the exon 7 of IRF6 gene, which encodes for the major region of protein binding domain. The PCR amplicons from all the samples were subsequently resolved in a 1.2% agarose gel, purified and subjected to direct sequencing to screen for mutations. Results: Sequencing analysis resulted in the identification of a mutation within exon 7 of IRF6 that occurred in heterozygous condition in 9% (3/32) of OSCC samples. The wild type codon TTC at position 252 coding for phenylalanine was found to be mutated to TAC that coded for tyrosine (F252Y). Conclusions: The present study identified for the first time a novel mutation within the conserved protein binding domain of IRF6 gene in tissue samples of subjects with OSCC.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 2002년도 창립10주년기념 및 국립독성연구원 의약품동등성평가부서 신설기념 국재학술대회:생물학적 동등성과 의약품 개발 전략을 위한 국제심포지움
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• pp.236-236
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• 2002

Pyruvate dehydrogenase phosphatase (PDP) is a mitochondrial protein serine/threonine phosphatase that catalyzes the dephosphorylation and concomitant reactivation of the pyruvate dehydrogenase componant of the pyruvate dehydrogenase complex (PDC). PDP consists of a Mg$\^$+2/ -dependent and Ca$\^$+2)-stimulated catalytic subunit (PDPc) of Mr 52,600 and a FAD-containing regulatory subunit (PDPr) of Mr 95.600. Catalytic subunit of pyruvate dehydrogenase phosphatase (PDPc) has been suggested to have three major functional domains such as dihydrolipoamide acetyltransferase(E$_2$)-binding domain, regulatory subunit of PDP(PDPr)-binding domain, and calcium-binding domain. In order to identify functional domains, recombinant catalytic subunit of pyruvate dehydrogenase phosphatase (rPDPc) was expressed in E. coli JM101 and purified to near homogeneity using the unique property of PDPc: PDPm binds to the inner lipoyl domain (L$_2$) of E$_2$ of pyruvate dehydrogenase complex (PDC) in the presence of Ca$\^$+2/, not under EGTA. PDPc was limited-proteolysed by trypsin, chymotrypsin, Arg-C, and elastase at pH7.0 and 30$^{\circ}C$ and N-terminal analysis of the fragment was done. Chymotrypsin, trypsin, and elastase made two major framents: N-terminal large fragment, approx. 50kD and C-terminal small fragment, approx. 0 kDa. Arg-C made three major fragments: N-terminal fragment, approx. 35 kD, and central fragment, approx. 15 kD, and C-terminal fragment, approx. 10 kD. This study strongly suggest that PDPc consists of three major functional domains. However, further study should be necessary to identify the functional role.

• 디지털융복합연구
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• 제15권12호
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• pp.25-33
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• 2017

이 연구에서는 교육현장, 상담 현장에서 실시되고 있는 정서지능 향상 프로그램의 효과에 대한 융합적 연구로서, 메타분석을 실시하였다. 메타분석은 통계적인 방법을 활용하여 선행연구물들에 대한 종합적인 결과와 프로그램 개입의 크기와 방향을 제시한다는 장점을 가지고 있는 분석방법이다. 논문의 선정기준을 통해 총 33편의 연구물을 분석 대상으로 선정하였다. 선정된 33편의 연구물에서 92개의 효과크기를 종합하여 분석한 결과 정서지능 향상 프로그램의 전체 효과크기는 0.839로 나타났다. 정서지능의 하위 범주 영역에 따른 결과를 비교한 결과, 정서인식 영역의 효과크기가 가장 큰 것으로 나타냈다(g=.919). 다음으로는 정서이입 영역(g=0.841), 정서활용 영역(g=0.834), 정서조절 영역(g=0.785), 정서표현 영역(g=0.766)순으로 효과크기를 보고하였다. 참가자의 연령에 따른 효과크기를 비교한 결과 대학생 및 성인, 초등학생, 중 고등학생, 순으로 효과크기가 나타났다. 이러한 연구 결과를 바탕으로 후속연구와 향후 프로그램 운영에 대한 시사점을 논의하였다.

• Earthquakes and Structures
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• 제17권2호
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• pp.143-162
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• 2019

Motivated by a simpler and more compact hybrid active tuned mass damper (ATMD) system with wide frequency spacing (i.e., high robustness) but not reducing the effectiveness using the least number of ATMD units, the active tuned tandem mass dampers (ATTMD) have been proposed to attenuate undesirable oscillations of structures under the ground acceleration. Likewise, it is expected that the frequency spacing of the ATTMD is comparable to that of the active multiple tuned mass dampers (AMTMD) or the multiple tuned mass dampers (MTMD). In accordance with the mode generalised system in the specific vibration mode being controlled (simply referred herein to as the structure), the closed-form expression of the dimensionless displacement variances has been derived for the structure with the attached ATTMD. The criterion for the optimum searching may then be determined as minimization of the dimensionless displacement variances. Employing the gradient-based optimization technique, the effects of varying key parameters on the performance of the ATTMD have been scrutinized in order to probe into its superiority. Meanwhile, for the purpose of a systematic comparison, the optimum results of two active tuned mass dampers (two ATMDs), two tuned mass dampers (two TMDs) without the linking damper, and the TTMD are included into consideration. Subsequent to work in the frequency domain, a real-time Simulink implementation of dynamic analysis of the structure with the ATTMD under earthquakes is carried out to verify the findings of effectiveness and stroke in the frequency domain. Results clearly show that the findings in the time domain support the ones in the frequency domain. The whole work demonstrates that ATTMD outperforms two ATMDs, two TMDs, and TTMD. Thereinto, a wide frequency spacing feature of the ATTMD is its highlight, thus deeming it a high robustness control device. Furthermore, the ATTMD system only needs the linking dashpot, thus embodying its simplicity.

• Journal of Microbiology and Biotechnology
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• 제29권5호
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• pp.813-819
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• 2019

Middle East respiratory syndrome coronavirus (MERS-CoV) induces severe respiratory impairment with a reported mortality rate of ~36% in humans. The absence of clinically available MERS-CoV vaccines and treatments to date has resulted in uncontrolled incidence and propagation of the virus. In vaccine design, fusion with the IgG Fc domain is reported to increase the immunogenicity of various vaccine antigens. However, limited reports have documented the potential negative effects of Fc fusion on vaccine antigens. To determine whether Fc fusion affects the immunogenicity of MERS-CoV antigen, we constructed a Fcassociated MERS-CoV spike protein (eS770-Fc, 110 kDa), whereby human IgG4 Fc domain was fused to MERS-CoV spike protein (eS770) via a Gly/Pro linker using baculovirus as the expression system. For comparative analyses, two eS770 proteins lacking the IgG4 Fc domain were generated using the IdeS protease ($eS770-{\Delta}Fc$) or His tag attachment (eS770-His) and the immunogenicity of the above constructs were examined following intramuscular immunization in mice. Contrary to expectations, non-Fc spike proteins ($eS770-{\Delta}Fc$, eS770-His; 90 kDa) showed higher immunogenicity than the Fc fusion protein (eS770-Fc). Moreover, unlike non-Fc spike proteins, eS770-Fc immunization did not elicit neutralizing antibodies against MERS-CoV. The lower immunogenicity of Fc-fused eS770 was related to alterations in the structural conformation of the spike protein. Taken together, our results indicate that IgG Fc fusion reduces the immunogenicity of eS770 by interfering with the proper folding structure.

• Journal of Microbiology and Biotechnology
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• 제33권7호
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• pp.964-972
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• 2023

Bacteriophage endolysins are peptidoglycan hydrolases composed of cell binding domain (CBD) and an enzymatically active domain. A phage endolysin CBD can be used for detecting bacteria owing to its high specificity and sensitivity toward the bacterial cell wall. We aimed to develop a method for detection of Enterococcus faecalis using an endolysin CBD. The gene encoding the CBD of ECP3 phage endolysin was cloned into the Escherichia coli expression vector pET21a. A recombinant protein with a C-terminal 6-His-tag (CBD) was expressed and purified using a His-trap column. CBD was adsorbed onto epoxy magnetic beads (eMBs). The bacterial species specificity and sensitivity of bacterial binding to CBD-eMB complexes were determined using the bacterial colony counting from the magnetic separations after the binding reaction between bacteria and CBD-eMB complexes. E. faecalis could bind to CBD-eMB complexes, but other bacteria (such as Enterococcus faecium, Staphylococcus aureus, Escherichia coli, Acinetobacter baumannii, Streptococcus mutans, and Porphyromonas gingivalis) could not. E. faecalis cells were fixed onto CBD-eMB complexes within 1 h, and >78% of viable E. faecalis cells were recovered. The E. faecalis recovery ratio was not affected by the other bacterial species. The detection limit of the CBD-eMB complex for E. faecalis was >17 CFU/ml. We developed a simple method for the specific detection of E. faecalis using bacteriophage endolysin CBD and MBs. This is the first study to determine that the C-terminal region of ECP3 phage endolysin is a highly specific binding site for E. faecalis among other bacterial species.

• 복식
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• 제52권4호
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• pp.155-171
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• 2002

Trompe-1’oeil technique strategically conceived with a view to effectively realize creative ideas among the expression techniques of fine arts style has provided the driving force in development of fine arts and has continuously influenced development of the modern fashion pursuing unique individuality. The purpose of this study is to open a new horizon for the development of fashion as a practical art, and to seek the expansion of the creative domain and ultimately to contribute to the creation of original and creative fashion by examining the interrelationship between Trompe-1’oeil, which has long been utilized and positioned as one of the leading fine arts techniques with the advent of surrealism in the beginning of the 20th century and the modern fashion. The study is focused first on finding out how Trompe-1’oeil technique originated in connection with researching the fashion of Trompe-1’oeil and on analysing the techniques of expression, and then on investigating into Elsa Schiaparelli, pioneer of Trompe-1’oeil technique to identify her influences, and finally on classifying clothes employing Trompe-1’oeil technique by their expression method to examine how Trompe-1’oeil technique has been applied to modern clothes. As for the research method, the researcher has referred to fine arts books, collection of pictorial records and the like to gain conceptual understanding of Trompe-1’oeil and to examine the expression method and the features of Trompe-1’oeil, and collected and referred to fashion books and fashion marazines to understand Elsa Schiaparelli and the expression tendencies of Tromprf-1’oeil in modern fashions. Particularly, the researcher has attempted to search the correlation between modern fashion and Trompe-1’oeil technique. As a result of this research, the researcher has managed to classify Trompe-1’oeil technique expressed in modern fashion into ‘harmony’, ‘application of the human body’, ‘front and back’, ‘surface and inside’. ‘completion of the incomplete’ and ‘detail.’ The researcher has also noted that Elsa Schiaparelli, a surrealist first applied Trompe-1’oeil technique to clothes and confirmed that quite a few avant-garde clothes designers following Elia Schiaparelli, by using Trompe-1’oeil technique in clothes, recently recreate fresh feelings.

• Molecules and Cells
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• 제27권1호
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• pp.75-81
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• 2009

The Arabidopsis gene AtLEC (At3g15356) gene encodes a putative 30-kDa protein with a legume lectin-like domain. Likely to classic legume lectin family of genes, AtLEC is expressed in rosette leaves, primary inflorescences, and roots, as observed in Northern blot analysis. The accumulation of AtLEC transcript is induced very rapidly, within 30 min, by chitin, a fungal wall-derived oligosaccharide elictor of the plant defense response. Transgenic Arabidopsis carrying an AtLEC promoter-driven ${\beta}$-glucuronidase (GUS) construct exhibited GUS activity in the leaf veins, secondary inflorescences, carpel heads, and silique receptacles, in which no expression could be seen in Northern blot analysis. This observation suggests that AtLEC expression is induced transiently and locally during developmental processes in the absence of an external signal such as chitin. In addition, mechanically wounded sites showed strong GUS activity, indicating that the AtLEC promoter responds to jasmonate. Indeed, methyl jasmonate and ethylene exposure induced AtLEC expression within 3-6 h. Thus, the gene appears to play a role in the jasmonate-/ethylene-responsive, in addition to the chitin-elicited, defense responses. However, chitin-induced AtLEC expression was also observed in jasmonate-insensitive (coi1) and ethylene-insensitive (etr1-1) Arabidopsis mutants. Thus, it appears that chitin promotes AtLEC expression via a jasmonate- and/or ethylene-independent pathway.

• Plant Biotechnology Reports
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• 제3권1호
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• pp.95-101
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• 2009

Tea leaves are major source of catechins—antioxidant flavonoids. Dihydroflavonol 4-reductase (DFR, EC 1.1.1.219) is one of the important enzymes that catalyzes the reduction of dihydroflavonols to leucoanthocyanins, a key ''late'' step in the biosynthesis of catechins. This manuscript reports characterization of DFR from tea (CsDFR) that comprised 1,413 bp full-length cDNA with ORF of 1,044 bp (115-1,158) and encoding a protein of 347 amino acids. Sequence comparison of CsDFR with earlier reported DFR sequences in a database indicated conservation of 69-87% among amino acid residues. In silico analysis revealed CsDFR to be a membrane-localized protein with a domain (between 16 and 218 amino acids) resembling the NAD-dependent epimerase/dehydratase family. The theoretical molecular weight and isoelectric point of the deduced amino sequence of CsDFR were 38.67 kDa and 6.22, respectively. Upon expression of CsDFR in E. coli, recombinant protein was found to be functional and showed specific activity of 42.85 nmol $min^{-1}$ mg $protein^{-1}$. Expression of CsDFR was maximum in younger rather than older leaves. Expression was down-regulated in response to drought stress and abscisic acid, unaffected by gibberellic acid treatment, but up-regulated in response to wounding, with concomitant modulation of catechins content. This is the first report of functionality of recombinant CsDFR and its expression in tea.