• Journal of Plant Biotechnology
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• 제30권3호
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• pp.281-291
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• 2003

In this review, we described the catalogues of the rice proteome which were constructed in our program, and functional characterization of some of these proteins was discussed. Mass-spectrometry is the most prevalent technique to rapidly identify a large number of proteome analysis. However, the conventional Western blotting/sequencing technique has been used in many laboratories. As a first step to efficiently construct protein cata-file in proteome analysis of major cereals, we have analyzed the N-terminal sequences of 100 rice embryo proteins and 70 wheat spike proteins separated by two-dimensional electrophoresis. Edman degradation revealed the N-terminal peptide sequences of only 31 rice proteins and 47 wheat proteins, suggesting that the rest of separated protein sports are N-terminally blocked. To efficiently determine the internal sequence of blocked proteins, we have developed a modified Cleveland peptide mapping method. Using this above method, the internal sequences of all blocked rice proteins(i, e., 69 proteins) were determined. Among these 100 rice proteins, thirty were proteins for which homologous sequence in the rice genome database could be identified. However, the rest of the proteins lacked homologous proteins. This appears to be consistent with the fact that about 45% of total rice cDNA have been deposited in the EMBL database. Also, the major proteins involved in the growth and development of rice can be identified using the proteome approach. Some of these proteins, including a calcium-binding protein that tuned out to be calreticulin, gibberellin-binding protein, which is ribulose-1.5-bisphosphate carboxylase/oxygense active in rice, and leginsulin-binding protein in soybean have functions in the signal transduction pathway. Proteomics is well suited not only to determine interaction between pairs of proteins, but also to identify multisubunit complexes. Currently, a protein-protein interaction database for plant proteins(http://genome.c.kanazawa-u.ac.jp/Y2H)could be a very useful tool for the plant research community. Also, the information thus obtained from the plant proteome would be helpful in predicting the function of the unknown proteins and would be useful be in the plant molecular breeding.

• Journal of Microbiology
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• 제34권1호
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• pp.25-29
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• 1996

Since sequencing of randomly selected cDNA clones has been known to be a powerful approach to obtain information on gene expression pattern in specific cells or tissues, we have analyzed a 3'-directed cDNA library of vegetative mycelia of A. nidulans by single-pass sequencing of hundreds of randomly selected clones. Sequencing of 292 cDNA clones yielded 209 gene signatures (GSs) probably representing highly or lesser expressed genes in the vegetative mycelia. Among the 209 GSs, 25 (79 cDNA clones) appeared more than once and 184 only once. One GS appeared at a highest frequency of 6 times, 2 GSs5 times, 4 GSs 4 times, a GSs 3 times and 16 GSs twice. About 6.6% GSs comprizing of 13 GSs showed alternative polyadenylation. Among 23 redundant GSs, three were common in both mycelia and sexual organs, and 22 were probably mycelia-specific. Out of 209 GSs, 36 were identified in GenBank showing of 70% or greater similaritis. Only six GSs were for A. nidulans genes, and 13 GSs were of DNA or genes encoding cytoplasmic or organellar proteins. This pattern is similar to those in the human HepG2 cell line and in human colonic mucosa, although very few genes for nuclear proteins and for protein synthesis were in A. nidulans.

• Journal of Plant Biotechnology
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• 제32권4호
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• pp.269-273
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• 2005

Methylotrophic 효모 Pichia pastoris 발현시스템을 사용하여 인간 TLR9 단백질의 세포내 TIR 도메인을 발현하였다. TIR 단백질이 P. pastoris에서 발현되어 배지 속으로 분비되는 것을 SDS-PAGE로 확인하였고, 발현된 단백질을 western-blot, MALDI-TOF 질량분석으로 동정하였다. 이를 통하여 TIR 딘백질이 P. pastoris에서 안정적으로 발현됨을 알 수 있었다. 그리고 발현된 단백질을 니켈 친화, 양이온교환수지, 겔 투과 크로마토그라피를 사용하여 순수 분리 정제하였다. P. pastoris를 이용한 단백질의 발현과 정제방법은 대장균에서 잘 발현되지 않는 단백질의 발현에 응용될 수 있을 것이다.

• 대한화장품학회:학술대회논문집
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• 대한화장품학회 2003년도 IFSCC Conference Proceeding Book I
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• pp.590-600
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• 2003

The matrix metalloproteinases (MMPs) are a family of enzymes responsible for degrading connective tissue. MMPs catalyze the breakdown of collagen from the extracellular matrix, leading to wrinkle formation and accelerated skin aging. Furthermore, ultraviolet irradiation causes increased expression of certain MMPs. In the extracellular matrix turnover, MMPs are interacting with endogenous regulators named tissue inhibitors of metalloproteinases (TIMPs). Using peptide substrate assays, it has been demonstrated that TIMP-MMP complexes interact highly specifically with $K_{i}$ values of 10$^{-9}$ -10$^{-16}$ M. Therefore applications for TIMP as inhibitor of collagen degradation are suggested for cosmetic anti-aging products to prevent wrinkle formation and loss of elasticity. To date four TIMP proteins (TIMP-1, TIMP-2, TIMP-3 and TIMP-4) have been identified which show a high degree in sequence similarity. The production of human TIMP-2, a 194-residue nonglycosylated protein, was performed by fed-batch culture of Escherichia coli. TIMP-2 accumulated in the bacterial cells in an insoluble form as inclusion bodies. The inclusion bodies were solubilized and the protein refolded to yield the native TIMP-2 in the active form. The integrity of the protein was confirmed by mass analysis, Edman sequencing and gel shift experiments with authentic samples. The inhibitory activity of the refolded and purified TIMP-2 was demonstrated with MMP-1 and MMP-2 assays using synthetic fluorogenic peptide substrates.s.

• Journal of Microbiology and Biotechnology
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• 제20권3호
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• pp.460-466
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• 2010

A mutant of green fluorescent protein ($GFPmut3^*$) from the jellyfish Aequorea victoria was cyclized in vitro and in vivo by the use of a naturally split intein from the dnaE gene of Synechocystis species PCC6803 (Ssp). Cyclization of $GFPmut3^*$ was confirmed by amino acid sequencing and resulted in an increased electrophoretic mobility compared with the linear $GFPmut3^*$. The circular $GFPmut3^*$ was $5^{\circ}C$ more thermostable than the linear form and significantly more resistant to proteolysis of exopeptidase. The circular $GFPmut3^*$ also displayed increased relative fluorescence intensity. In addition, chemical stability of $GFPmut3^*$ against GdnHCl revealed more stability of the circular form compared with the linear form.

• Journal of Animal Science and Technology
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• 제62권2호
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• pp.121-140
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• 2020

Anaerobic fungi habitat in the gastrointestinal tract of foregut fermenters or hindgut fermenters and degrade fibrous plant biomass through the hydrolysis reactions with a wide variety of cellulolytic enzymes and physical penetration through fiber matrix with their rhizoids. To date, seventeen genera have been described in family Neocallimasticaceae, class Neocallimastigomycetes, phylum Neocallimastigomycota and one genus has been described in phylum Neocallimastigomycota. In National Center for Biotechnology Information (NCBI) database (DB), 23,830 nucleotide sequences and 59,512 protein sequences have been deposited and most of them were originated from Piromyces, Neocallimastix and Anaeromyces. Most of protein sequences (44,025) were acquired with PacBio next generation sequencing system. The whole genome sequences of Anaeromyces robustus, Neocallimastix californiae, Pecoramyces ruminantium, Piromyces finnis and Piromyces sp. E2 are available in Joint Genome Institute (JGI) database. According to the results of protein prediction, average Isoelectric points (pIs) were ranged from 5.88 (Anaeromyces) to 6.57 (Piromyces) and average molecular weights were ranged from 38.7 kDa (Orpinomyces) to 56.6 kDa (Piromyces). In Carbohydrate-Active enZYmes (CAZY) database, glycoside hydrolases (36), carbohydrate binding module (11), carbohydrate esterases (8), glycosyltransferase (5) and polysaccharide lyases (3) from anaerobic fungi were registered. During four decades, 1,031 research articles about anaerobic fungi were published and 444 and 719 articles were available in PubMed (PM) and PubMed Central (PMC) DB.

• Journal of Microbiology and Biotechnology
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• 제10권5호
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• pp.670-676
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• 2000

Several clones obtained from Serratia marcescens stimulated E. coli TP2139 (${\Delta}lac, \;{\Delta} crp$) cells to use maltose as a carbon source. The crp gene clone, pCKB12, was confirmed to stimulate the $\beta$-galactosidase activity, by Southern hybridization [31]. The nucleotide sequence of the crp region consisting of 1,979 bp was determined. The sequencing of the fragment led to the identification of two open reading frames: One of these, the crp gene, encoded 210 amino acid and the other encoded a truncated protein. The S. marcescens and E. coli crp genes showed a higher degree of divergence in their nucleotide sequence with 120 changes, however, the corresponding amino acid sequences showed only two amino acid differences. Yet, an analysis of the amino acid divergence revealed that the catabolite gene activator protein, the crp gene product, was the most conserved protein observed so far. Using a crp-lac protein fusion, it was demonstrated that S. marcescens CRP could repress its own expression, probably via a mechanism similar to that previously described for the E. coli crp gene.

• 한국미생물·생명공학회지
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• 제23권6호
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• pp.678-686
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• 1995

SecY is a central component of the protein export machinery that mediate the translocation of secretory proteins across the plasma membrane of Escherichia coli. In order to study the mechanism of protein secretion in Streptomyces, we have done cloning and sequencing of the Streptomyces coelicolor secY gene by using polymerase chain reaction method. The nucleotide sequence of the gene for SecY from S. coelicolor showed over 58% identity to that of M. luteus. The deduced amino acid sequences were highly homologous to those of other known SecY polypeptides, all having the potential to form 10 transmembrane segments, and especially second, fifth, and tenth segments were particularly conserved, sharing greater than 75% identity with W. lute s SecY. We propose that the conserved membrane-spanning segments actively participate in protein export. In B. subtilis and E. coli, the secY gene is a part of the spc operon, is preceded by the gene coding for ribosomal protein L15, and is likety coupled transcriptionally and translationally to the upstream L15 gene. In the other hand, secY gene of S. coelicolor and M. luteus have its own promoter region, are coupled translationally with adk gene and pr sented in adk operon.

• Genomics & Informatics
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• 제19권2호
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• pp.14.1-14.6
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• 2021

Coronavirus disease 2019 (COVID-19) is an on-going pandemic disease infecting millions of people across the globe. Recent reports of reduction in antibody levels and the re-emergence of the disease in recovered patients necessitated the understanding of the pandemic at the core level. The cases of multiple organ failures emphasized the consideration of different organ systems while managing the disease. The present study employed RNA sequencing data to determine the disease associated differentially regulated genes and their related protein interactions in several organ systems. It signified the importance of early diagnosis and treatment of the disease. A map of protein interactions of multiple organ systems was built and uncovered CAV1 and CTNNB1 as the top degree nodes. A core interactions sub-network was analyzed to identify different modules of functional significance. AR, CTNNB1, CAV1, and PIK3R1 proteins were unfolded as bridging nodes interconnecting different modules for the information flow across several pathways. The present study also highlighted some of the druggable targets to analyze in drug re-purposing strategies against the COVID-19 pandemic. Therefore, the protein interactions map and the modular interactions of the differentially regulated genes in the multiple organ systems would incline the scientists and researchers to investigate in novel therapeutics for the COVID-19 pandemic expeditiously.

• BMB Reports
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• 제52권5호
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• pp.304-311
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• 2019

The cytoplasmic FMR1-interacting protein family (CYFIP1 and CYFIP2) are evolutionarily conserved proteins originally identified as binding partners of the fragile X mental retardation protein (FMRP), a messenger RNA (mRNA)-binding protein whose loss causes the fragile X syndrome. Moreover, CYFIP is a key component of the heteropentameric WAVE regulatory complex (WRC), a critical regulator of neuronal actin dynamics. Therefore, CYFIP may play key roles in regulating both mRNA translation and actin polymerization, which are critically involved in proper neuronal development and function. Nevertheless, compared to CYFIP1, neuronal function and dysfunction of CYFIP2 remain largely unknown, possibly due to the relatively less well established association between CYFIP2 and brain disorders. Despite high amino acid sequence homology between CYFIP1 and CYFIP2, several in vitro and animal model studies have suggested that CYFIP2 has some unique neuronal functions distinct from those of CYFIP1. Furthermore, recent whole-exome sequencing studies identified de novo hot spot variants of CYFIP2 in patients with early infantile epileptic encephalopathy (EIEE), clearly implicating CYFIP2 dysfunction in neurological disorders. In this review, we highlight these recent investigations into the neuronal function and dysfunction of CYFIP2, and also discuss several key questions remaining about this intriguing neuronal protein.