• Applied Microscopy
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• v.52
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• pp.11.1-11.9
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• 2022

Zoonotic poxvirus infections pose significant threat to human health as we have witnessed recent spread of monkeypox. Therefore, insights into molecular mechanism behind poxvirus replication cycle are needed for the development of efficient antiviral strategies. Virion assembly is one of the key steps that determine the fate of replicating poxviruses. However, in-depth understanding of poxvirus assembly is challenging due to the complex nature of multi-step morphogenesis and heterogeneous virion structures. Despite these challenges, decades of research have revealed virion morphologies at various maturation stages, critical protein components and interactions with host cell compartments. Transmission electron microscopy has been employed as an indispensable tool for the examination of virion morphology, and more recently for the structure determination of protein complexes. In this review, we describe some of the major findings in poxvirus morphogenesis and the contributions of continuously advancing electron microscopy techniques.

• Archives of Pharmacal Research
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• v.20 no.5
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• pp.459-464
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• 1997

The human liver cDNA clone UDPGTh2, encoding a liver UDP-glucuronosyltransferase (UDPGT) was isolated from a .gamma. gt 11 cDNA library by hybridization to mouse transferase cDNA clone, UDPGTm1. UDPGTh2 encoded a 529 amino acid protein with an amino terminus membrane-insertion signal peptide and a carboxyl terminus membrane-spanning region. There were three potential asparagine-linked glycosylation sites at residues 67, 68, and 315. In order to obtain UDPGTh2 protein encoded from cloned human liver UDP-glucuronosyltransferase cDNA, the clone was inserted into the pSVL vector (pUDPGTh2) and expressed in COS 1 cells. The presence of a transferase with Mr-52,000 in transfected cells cultured in the presence of $[^{35}S]$ methionine was shown by immunocomplexed products with goat antimouse transferase IgG and protein A-Sepharose and analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. The expressed UDPGT was a glycoprotein as indicated by electrophoretic mobility shift in Mr-3,000-4,000 when expressed in the presence of tunicamycin. The extent of glycosylation was difficult to assess, although one could assume that glycosyl structures incorporated at the level of endoplasmic reticulum were always the core oligosaccharides. Thus, it is likely that at least two moieties inserted can account for the shift of Mr-3,000-4,000. This study demonstrates the cDNA and deduced amino acid sequence of human liver UDP-glucuronosyltransferase cDNA, UDPGTh2.

• Journal of Applied Biological Chemistry
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• v.50 no.1
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• pp.1-5
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• 2007

Chemical properties of spray-dried powders separated based on molecular weight from crude protein binding polysaccharide (CP-SD) of Agraricus blazei were examined. Contents of ${\beta}$-glucan in SD-1, SD-2 and SD-3 were 18.67%, 48.24%, and 37.15% respectively, and SD-2 (10-150 kDa) showed the highest molecular weight. Obtained ${\beta}$-glucans were not pure glucan, but was determined to be an acidic proteo-heteroglycan with a large amount of glucose (74.46-80.05%), galactose (8.91-15.2%), and mannose (4.9-5.46%). Composition of their amino acids was mainly aspartic and glutamic acids. FT-IR spectrum revealed SD-1, SD-2 and SD-3 were structures of ${\beta}$-1,3-glucans and ${\alpha}$-1,6-glucans at 890 and 930 $cm^{-1}$, respectively, signals of ${\alpha}$-1,6-glucans for CP-SD was not found. Useful CP-SD was recovered from A. blazei for preparation of three powder types as food materials.

• Korean Journal of Food Science and Technology
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• v.20 no.1
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• pp.72-78
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• 1988

Microstructures of 18 kinds of legume seeds (15 varieties including 3 strains) which were cultivated in tropical areas, were observed under a light microscope. Majority of legume seeds were composed of starchy cotyledonary cells in which large amounts of single starch granules were contained, while a few had cotyledonary cells filled with a number of protain bodies. Starch granules were different in size and shape depending on varieties. Some contained lipid bodies distributed in cytoplasmic network, and were distinctive in thick cell walls. Microstructure of soybean was also observed for the comparison of the structures.

• Biomedical Science Letters
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• v.18 no.2
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• pp.160-164
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• 2012

The baculovirus/insect cell expression system is of great value for the large-scale production of normal and mutant mammalian passive glucose-transport proteins heterologously for structural and functional studies. In most mammalian cells that express HepG2, this transporter isoform is predominantly located at the cell surface. However, it had been reported that heterologous expression of other membrane proteins using the baculovirus system induced highly vacuolated cytoplasmic membranes. Therefore, how a cell responds to the synthesis of large amounts of a glycoprotein could be an interesting area for investigation. In order to examine the subcellular location of the human HepG2 transport proteins when expressed in insect cells, immunofluorescence studies were carried out. Insect cells were infected with the recombinant baculovirus AcNPVHIS-GT or with wild-type virus at a MOI of 5, or were not exposed to viral infection. A high level of fluorescence displayed in cells infected with the recombinant virus indicated that transporters are expressed abundantly and present on the surface of infected Sf21 cells. The evidence for the specificity of the immunostaining was strengthened by the negative results shown in the negative controls. Distribution of the transporter protein expressed in insect cells was further revealed by making a series of optical sections through an AcNPVHIS-GT-infected cell using a confocal microscope, which permits optical sectioning of cell sample. These sections displayed intense cytoplasmic immunofluorecence surrounding the region occupied by the enlarged nucleus, indicating that the expressed protein was present not only at the cell surface but also throughout the cytoplasmic membranous structures.

• Journal of Microbiology and Biotechnology
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• v.25 no.7
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• pp.963-977
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• 2015

The well-characterized gram-positive bacterium Bacillus subtilis is an outstanding industrial candidate for protein expression owing to its single membrane and high capacity of secretion, simplifying the downstream processing of secretory proteins. During the last few years, there has been continuous progress in the illustration of secretion mechanisms and application of this robust host in various fields of life science, such as enzyme production, feed additives, and food and pharmaceutical industries. Here, we review the developments of Bacillus subtilis as a highly promising expression system illuminating strong chemical- and temperatureinducible and other types of promoters, strategies for ribosome-binding-site utilization, and the novel approach of signal peptide selection. Furthermore, we outline the main steps of the Sec pathway and the relevant elements as well as their interactions. In addition, we introduce the latest discoveries of Tat-related complex structures and functions and the countless applications of this full-folded protein secretion pathway. This review also lists some of the current understandings of ATP-binding cassette transporters. According to the extensive knowledge on the genetic modification strategies and molecular biology of Bacillus subtilis, we propose some suggestions and strategies for improving the yield of intended productions. We expect this to promote striking future developments in the optimization and application of this bacterium.

• Proceedings of the PSK Conference
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• 2002.10a
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• pp.315.3-316
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• 2002

A universal set of genes encodes the components of dissociated. type II. fa11y acid synthase system that is responsible for producing the multitude of fa11y acid structures found in bacterial membranes. We examined the biochemical basis for the production of fatty acids by bacteria. Several genes from HaemophHus influenzae Rd and three genes from Enterococcus faecalis V583 were predicted to encode homologs of the ${\beta}$-ketoacyl-acyl carrier protein synthases I or II or III of Escherichia coli(FabB or BabF, or FabH)were identified in the genomic database. The protein products were expressed. purified, and biochemically characterized. efFabH and hF abH carried out the initial condensation reaction of fatty acid biosynthesis with acetyl-Coenzyme A as a primer. and hFabB and efFabF1 carried out the elongation condensation reaction of fatty acid biosynthesis with myrixtoyl-ACP.

• Journal of Microbiology and Biotechnology
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• v.25 no.1
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• pp.127-136
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• 2015

Cupredoxin-like proteins are mainly copper-binding proteins that conserve a typical rigid Greek-key arrangement consisting of an eight-stranded β-sandwich, even though they share as little as 10-15% sequence similarity. The electron transport function of the Cupredoxins is critical for respiration and photosynthesis, and the proteins have therapeutic potential. Despite their crucial biological functions, the identification of the distant Cupredoxin homologs has been a difficult task due to their low sequence identity. In this study, the overlapped conserved residue (OCR) fingerprint for the Cupredoxin superfamily, which consists of conserved residues in three aspects (i.e., the sequence, structure, and intramolecular interaction), was used to detect the novel Cupredoxin homologs in the NCBI non-redundant protein sequence database. The OCR fingerprint could identify 54 potential Cupredoxin sequences, which were validated by scanning them against the conserved Cupredoxin motif near the Cu-binding site. This study also attempted to model the 3D structures and to predict the functions of the identified potential Cupredoxins. This study suggests that the OCR-based approach can be used efficiently to detect novel homologous proteins with low sequence identity, such as Cupredoxins.

• Korean Journal of Veterinary Research
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• v.47 no.2
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• pp.169-174
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• 2007

To provide information for the molecular pathogenesis and antigenic structures of Korean isolates of porcine epidemic diarrhea virus (PEDV), the small membrane (sM) protein gene of Chinju99 strain, which was previously isolated from piglets suffering from severe diarrhea was characterized and further analyzed with other PEDV strains. The sM gene of Chinju99 generated by reverse transcription and polymerase chain reaction had a single open reading frame with 231 bases consisting of 24.2% adenine, 18.6% cytosine, 18.1% guanine and 39.0% thymine nucleotides. Nucleotide sequence of the gene revealed 97.8% homology to those of Belgian strain CV777 and British strain Br1/87, and 97.0% to Chinese strain LZC. The gene encoded a protein with 76 amino acids, and putative amino acid sequence of the gene revealed 98.7% homology to those of CV777 and Br1/87, and 96.1% to LZC. The amino acids of Chinju99 sM gene consisted of mostly hydrophobic residues, and there were one potential N-myristylation site and one potential threonine (T)-linked phosphorylation site recognized. Also, there was a transmembrane region with 46 amino acids, and Chinju99 was more close to CV777 and Br1/87 than to LZC in phylogenetic analysis on the sM amino acid sequences.

• Applied Biological Chemistry
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• v.32 no.1
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• pp.64-72
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• 1989

Near full length cDNA clones encoding the rice seed storage protein, prolamine, were isolated and divided into two homology classes based on cross-hybridization and DNA sequencing analysis. These cDNA clones contain a single open reading frame encoding a putative rice prolamine precursor(M.W.=17,200) possessing atypical 14 amino acid signal peptide. Clones of these two homology classes diverge mainly by insertions/deletions of short nucleotide stretches and point mutations. The deduced primary structures of both types of prolamine polypeptides are devoid of any major tandem repetitive sequences, a feature prevalent in other cereal prolamines. No significant homology teas detected between the rice prolamine and other cereal prolamines, indicating that the rice gene evolved from a different ancestor that gave rise to other cereal prolamine genes. Developing wheat and rice endosperms were examined using ultrathin sections prepared from tissues harvested at various days after flowering. By immunocytochemical localization techniques, wheat prolamines are localized within vesicles from Golgi apparatus and in homogeneous regions of protein bodies. The involvement of the goli apparatus in the packaging of wheat prolamines into protein bodies indicates a pathway which differs from the mode of other cereal prolamines and resembles the mechanism employed for the storage of rice glutelin and legume globulins.