• Bulletin of the Korean Chemical Society
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• v.12 no.6
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• pp.714-716
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• 1991

• BMB Reports
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• v.40 no.5
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• pp.617-624
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• 2007

Protein arginine methylation is a posttranslational modification involved in various cellular functions including cell signaling, protein subcellular localization and transcriptional regulation. We analyze the protein arginine methyltransferases (PRMTs) that catalyze the formation of methylarginines in porcine brain. We fractionated the brain extracts and determined the PRMT activities as well as the distribution of different PRMT proteins in subcellular fractions of porcine brain. The majority of the type I methyltransferase activities that catalyze the formation of asymmetric dimethylarginines was in the cytosolic S3 fraction. High specific activity of the methyltransferase was detected in the S4 fraction (high-salt stripping of the ultracentrifugation precipitant P3 fraction), indicating that part of the PRMT was peripherally associated with membrane and ribosomal fractions. The amount and distribution of PRMT1 are consistent with the catalytic activity. The elution patterns from gel filtration and anion exchange chromatography also indicate that the type I activity in S3 and S4 are mostly from PRMT1. Our results suggest that part of the type I arginine methyltransferases in brains, mainly PRMT1, are sequestered in an inactive form as they associated with membranes or large subcellular complexes. Our biochemical analyses confirmed the complex distribution of different PRMTs and implicate their regulation and catalytic activities in brain.

• YAKHAK HOEJI
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• v.43 no.6
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• pp.709-715
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• 1999

Polyphenol oxidase (PPO) activity in 200×g (cell wall), 4,000×g (plastid), 100,000×g (mitochondrial) and soluble fractions of the perilla leaves was monitored in the upper, middle and lower sections of the plant. In the course of plant growth, PPO activities in plastid and mitochondrial fractions were decreased, while those in cell wall fraction were maintained. During growing process, specific activities and PPO activities of each fraction were decreased, while total phenol content were decreased in middle (middle) and then increased in later stage (lower). Cell wall, plastid, mitochondrial (pellet) and soluble fraction had slightly different pH optima and substrate specificities. Isoenzyme patterns were identical in two bands for PPO activity in different subcellular fractions. Their molecular weights were 37KD and 48KD respectively.

• 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.

• Korean Journal of Microbiology
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• v.42 no.4
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• pp.271-276
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• 2006

Previously we demonstrated that virus-inducible stress protein (VISP) is induced in fish cells by the infection of a fish rhabdovirus. In this paper, we investigated the subcellular localization of the VISP and determined the region of VISP responsible for the subcellular localization. The CHSE-214 cells were stained with monoclonal antibody raised against VISP and observed with confocal microscope to detect the endogenous VISP. The results showed that the VISP localizes to the perinuclear region as spots. A plasmid expressing VISP fused to enhanced green fluorescent protein (EGFP) was constructed. The transient expression of full-length VISP fused to EGFP in CHSE-214 cells confirmed the spot formation of the VISP at perinuclear region. To determine the region responsible for the perinuclear localization of the VISP, we constructed a series of deletion mutants and, by using these deletion mutants, we found that C-terminal region of the VISP (aa 612-710) is essential for the perinuclear distribution of VISP and that this region contained nuclear receptor binding motif (691-TLTSLLL-697). Our results suggest that VISP localizes to the perinuclear region and C-terminal regions are important for this localization. Further studies on the role of the perinuclear localization of VISP in IHNV growth mali reveal the novel mechanism of IHNV pathogenecity.

• Journal of Life Science
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• v.9 no.6
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• pp.722-727
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• 1999

We carried out immunoblot analyses to study expression and subcellular distribution of the N-methyl-D-aspartate receptor(NR) subunits in salmon (Chum Salmon, Oncorhynchus keta). We prepared subcellular fractions such as brain homogenates, synaptosomes, and postsynaptic density (PSD) from salmon brains, and analyzed protein compositions by SDS-polyacrylamide gel electrophoresis (SDS-PAGE). In a Coomassie-stained 6% SDS-gel, about 20 distinct major protein bands could be identified in the PSD fraction. Immunoblot analyses using antibodies against rat NR subunit 2A and 2B antigens (NR2A and NR2B, respectively) showed weak but evident signals at the 180 kDa positions in the salmon PSD fractions. However, in contrast to rat NRs, the salmon NR2A and NR2B are not recognized by a phosphotyrosine-specific antibody suggesting that the salmon NRs are regulated differently from those of the rat by protein tyrosine kinases. Our results indicate that NR2A and NR2B subunits are expressed in the salmon PSD fraction but not regulated by tyrosine phosphorylation.

• Korean Journal of Veterinary Research
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• v.39 no.1
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• pp.45-54
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• 1999

In the present study we have analyzed the characteristics and distribution of the mu-opioid receptor(MOR) by raising anti-peptide antisera to the C-terminal peptide of MOR. The antisera against MOR was produced in New Zealand White rabbit against 15 residue corresponding to amino acids, 384-398 of the cloned rat MOR. The antigenic peptide was synthesized using an Applied Biosystems 432 solid-phase peptide synthesizer. The specificity and identification of the antisera were tested by analysis of transfected cells, epitope mapping and immunohistochemical method. COS-7 cells electroporated with MOR cDNA were used to evaluate the characteristics and subcellular distribution of MOR. MOR immunoreactivity was prodominent in the plasmalemma and subcellular compartments such as endoplasmic reticulum, Golgi apparatus and vesicle like structure. Furthermore, both tissue sections and transfected cell lines could be immunostained with these antisera and the immunoreactivity was abolished when anti-MOR sera were preincubated with the peptide against which they were raised. Based on epitope mapping analysis, all antisera appeared to have a similar epitope, which was determined to be within the last amino acid, 391-398. Moreover, immunohistochemistry showed that MOR immunoreactivity was observed in many brain areas including cerebral cortex, striatum, hippocampus, locus coeruleus and the superficial laminae of the dorsal horn. These stained spinal cord and brain areas showed the mirrored pattern observed in auto radiographic studies of mu-opioid binding as well as a pattern similar to that seen by is situ hybridization for MOR. Thus, several lines of evidence support the conclusion that the antisera produced in the present study most likely recognize mu-opioid receptor. These results suggest that MOR antisera may be utilized as useful tool to analyze the physiological and pharmacological studies for mu-opioid receptor in the future.

• Parasites, Hosts and Diseases
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• v.21 no.1
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• pp.41-48
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• 1983

The activity and distribution of aspartate aminotransferase (EC 2.6. 1. 1) and alanine aminotransferase (EC 2.6.1.2) in adult Fascicle hepatica have been studied. Fasciola hepatica was fractionated by differential centrifugation into nuclear, mitochondrial and cytosolic fractions. The activity of GOT and GPT was measured by the method of Reitman and Frankel. Isozyme patterns of those enzyme were also examined by DEAE-cellulose column chromatography. The results obtained were as follows; 1. The activity of aspartate and alanine aminotransferase was about 0.55 unit and 0.92 unit per 1g of Fascicle hepatica, respectively. 2. The activity of those enzymes was relatively low compared with those in mammalian tissues. 3. The distribution of aspartate aminotransferase in the subcellular organelles showed that 71% of the activity was in cytosolic, 24% in mitochondrial and 5% was in nuclear fraction. 4. About 22% of the total alanine aminotransferase activity was found in the mitochondrial fratstion, about 66% in the cytosolic fraction. 5. Aspartate aminotransferase from cytosolic fraction was separated into two types of isozymes, whereas alanine aminotransferase from cytosolic fraction gave only one active peak on DEAE-cellulose column chromatography.

• Molecules and Cells
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• v.22 no.1
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• pp.97-103
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• 2006

Phosphoinositides are critical regulators of ion channel and transporter activity. There are multiple isomers of biologically active phosphoinositides in the plasma membrane and the different lipid species are non-randomly distributed. However, the mechanism by which cells impose selectivity and directionality on lipid movements and so generate a non-random lipid distribution remains unclear. In the present study we investigated which structural elements of phosphoinositides are responsible for their subcellular location and movement. We incubated phosphatidylinositol (PI), phosphatidylinositol 4-monophosphate (PI(4)P) and phosphatidylinositol 4,5-bisphosphate ($PI(4,5)P_2$) with short or long acyl chains in CHO and HEK cells. We show that phosphate number and acyl chain length determine cellular location and translocation movement. In CHO cells, $PI(4,5)P_2$ with a long acyl chain was released into the cytosol easily because of a low partition coefficient whereas long chain PI was released more slowly because of a high partition coefficient. In HEK cells, the cellular location and translocation movement of PI were similar to those of PI in CHO cells, whereas those of $PI(4,5)P_2$ were different; some mechanism restricted the translocation movement of $PI(4,5)P_2$, and this is in good agreement with the extremely low lateral diffusion of $PI(4,5)P_2$. In contrast to the dependence on the number of phosphates of the phospholipid head group of long acyl chain analogs, short acyl chain phospholipids easily undergo translocation movement regardless of cell type and number of phosphates in the lipid headgroup.

• Geophysics and Geophysical Exploration
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• v.4 no.2
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• pp.45-54
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• 2001

A three-dimensional finite difference time-domain modeling algorithm based on staggered grid and considering transmitting and receiving antennas has been developed to simulate Ground Penetrating Radar (GPR) survey. This algorithm adopted the subcellular method to simulate the dipole antennas being used in GPR system and added resistors to reduce ringing caused by the reflections at the ends of an antenna. Comparison of the output voltages in the presence of the resistors for half-space said that the ringing and the amplitude of output voltage decreased as the number of resistors increased, and the antenna was designed based upon this result. Radiation patterns were derived to understand the distribution of electric field energy in the planes including or normal to the antenna. The electric field energy concentrated on vertical direction in the plane including antenna more than in normal plane.