• Journal of Microbiology and Biotechnology
• /
• v.17 no.2
• /
• pp.207-217
• /
• 2007

The Saccharomyces0 cerevisiae KNU5377 strain, which was isolated from spoilage in nature, has the ability to convert biomass to alcohol at high temperatures and it can resist against various stresses [18, 19]. In order to understand the defense mechanisms of the KNU5377 strain under menadione (MD) as oxidative stress, we used several techniques for study: peptide mass fingerprinting (PMF) by matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (MS) followed by two-dimensional (2D) gel electrophoresis, liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS), and surface-enhanced laser desorption ionization-time of flight (SELDI-TOF) technology. Among the 35 proteins identified by MALDI-TOF MS, 19 proteins including Sod1p, Sod2p, Tsa1p, and Ahp1p were induced under stress condition, while 16 proteins were augmented under normal condition. In particular, five proteins, Sod1p, Sod2p, Ahp1p, Rib3p, Yaf9p, and Mnt1p, were induced in only stressed cells. By LC-ESI-MS/MS analysis, 37 proteins were identified in normal cells and 49 proteins were confirmed in the stressed cells. Among the identified proteins, 32 proteins were found in both cells. Five proteins including Yel047cp and Met6p were only upregulated in the normal cells, whereas 17 proteins including Abp1P and Sam1p were elevated in the stressed cells. It was interesting that highly hypothetical proteins such as Ynl281wp, Ygr279cp, Ypl273wp, Ykl133cp, and Ykr074wp were only expressed in the stressed cells. SELDI-TOF analysis using the SAX2 and WCX2 chips showed that highly multiple-specific protein patterns were reproducibly detected in ranges from 2.9 to 27.0 kDa both under normal and stress conditions. Therefore, induction of antioxidant proteins, hypothetical proteins, and low molecular weight proteins were revealed by different proteomic techniques. These results suggest that comparative analyses using proteomics might contribute to elucidate the defense mechanisms of KNU5377 under MD stress.

• Mass Spectrometry Letters
• /
• v.13 no.3
• /
• pp.84-94
• /
• 2022

Neoadjuvant chemoradiotherapy (nCRT) is a standard therapy used for locally advanced rectal cancer prior to surgery, which can more effectively reduce the locoregional recurrence rate and radiation toxicity compared to postoperative chemoradiotherapy. The response of patients to nCRT varies, and thus, robust biomarkers for predicting a pathological complete response are necessary. This study aimed to identify possible biomarkers involved in the complete response/non-response of rectal cancer patients to nCRT. Comparative proteomic analysis was performed on rectal tissue samples before and after nCRT. Proteins were extracted for label-free proteomic analysis. Western blot and real-time PCR were performed using rectal cancer cell line SNU-503 and radiation-resistant rectal cancer cell line SNU-503R80Gy. A total of 135 up- and 93 down-regulated proteins were identified in the complete response group. Six possible biomarkers were selected to evaluate the expression of proteins and mRNA in SNU-503 and SNU-503R80Gy cell lines. Lyso-phosphatidylcholine acyltransferase 2, annexin A13, aldo-ketose reductase family 1 member B1, and cathelicidin antimicrobial peptide appeared to be potential biomarkers for predicting a pathological complete response to nCRT. This study identified differentially expressed proteins and some potential biomarkers in the complete response group, which would be further validated in future studies.

• Korean Journal of Oriental Medicine
• /
• v.13 no.1 s.19
• /
• pp.153-160
• /
• 2007

In the post-genome era, analysis of the cellular transcriptome using microarray or the cellular proteome using a 2-D gel electrophoresis and MALDI-TOF mass spectrometry are most widely used. Stroke is one of the most important causes of death along with cancer and cardiac disease. When pathological change of cells in developed from cerebral ischemia accompanied by stroke administration of neuroprotective drugs before stroke can decreases the degeneration of neuronal cells. The purpose of the present study was to assess the neuroprotective effect and protein expression after administration of P004, middle cerebral artery model of cerebral ischemia in rats. SD rats were subjected to middle cerebral artery occlusion. P004 (1,000 mg/kg) was administered 2 times at 0, 90 minutes after middle cerebral artery occlusion (MCAo). Rats were killed at 48 hours, and infarct area and volume were determined by histology and computerized image analysis. We investigated the protein expression profile on the global ischemia induced by MCAo. This proteomic analysis enable us to identify several proteins differently expressed in infarct brain tissue. The aims of this study were to do investigation comparing the neuroprotection activities of P004 and to understand the mechanism of acted as neuroprotective drug.

• Journal of Microbiology and Biotechnology
• /
• v.18 no.8
• /
• pp.1368-1376
• /
• 2008

In our previous study, the expression of active H-ferritins in Saccharomyces cerevisiae was found to reduce cell growth and reactive oxygen species (ROS) generation upon exposure to oxidative stress; such expression enhanced that of high-affinity iron transport genes (FET3 and FTR1). The results suggested that the recombinant cells expressing H-ferritins induced cytosolic iron depletion. The present study analyzes metabolic changes under these circumstances via proteomic methods. The YGH2 yeast strain expressing A-ferritin, the YGH2-KG (E62K and H65G) mutant strain, and the YGT control strain were used. Comparative proteomic analysis showed that the synthesis of 34 proteins was at least stimulated in YGH2, whereas the other 37 proteins were repressed. Among these, the 31 major protein spots were analyzed via nano-LC/MS/MS. The increased proteins included major heat-shock proteins and proteins related to endoplasmic reticulum-associated degradation (ERAD). On the other hand, the proteins involved with folate metabolism, purine and methionine biosynthesis, and translation were reduced. In addition, we analyzed the insoluble protein fractions and identified the fragments of Idh1p and Pgk1p, as well as several ribosomal assembly-related proteins. This suggests that intracellular iron depletion induces imperfect translation of proteins. Although the proteins identified above result from changes in iron metabolism (i.e., iron deficiency), definitive evidence for iron-related proteins remains insufficient. Nevertheless, this study is the first to present a molecular model for iron deficiency, and the results may provide valuable information on the regulatory network of iron metabolism.

• Journal of Microbiology and Biotechnology
• /
• v.29 no.11
• /
• pp.1817-1829
• /
• 2019

Porcine deltacoronavirus (PDCoV) is an emerging swine enteric coronavirus that causes diarrhea in neonatal piglets. Like other coronaviruses, PDCoV encodes at least three accessory or species-specific proteins; however, the biological roles of these proteins in PDCoV replication remain undetermined. As a first step toward understanding the biology of the PDCoV accessory proteins, we established a stable porcine cell line constitutively expressing the PDCoV NS7 protein in order to investigate the functional characteristics of NS7 for viral replication. Confocal microscopy and subcellular fractionation revealed that the NS7 protein was extensively distributed in the mitochondria. Proteomic analysis was then conducted to assess the expression dynamics of the host proteins in the PDCoV NS7-expressing cells. High-resolution two-dimensional gel electrophoresis initially identified 48 protein spots which were differentially expressed in the presence of NS7. Seven of these spots, including two up-regulated and five down-regulated protein spots, showed statistically significant alterations, and were selected for subsequent protein identification. The affected cellular proteins identified in this study were classified into functional groups involved in various cellular processes such as cytoskeleton networks and cell communication, metabolism, and protein biosynthesis. A substantial down-regulation of α-actinin-4 was confirmed in NS7-expressing and PDCoV-infected cells. These proteomic data will provide insights into the understanding of specific cellular responses to the accessory protein during PDCoV infection.

• Asian Pacific Journal of Cancer Prevention
• /
• v.13 no.12
• /
• pp.6435-6439
• /
• 2012

Chemoresistance to cancer therapy is a major obstacle to the effective treatment of human cancers with cisplatin (DDP), but the mechanisms of cisplatin-resistance are not clear. In this study, we established a cisplatin-resistant human ovarian cancer cell line (COC1/DDP) and identified differentially expressed proteins related to cisplatin resistance. The proteomic expression profiles in COC1 before and after DDP treatment were examined using 2-dimensional electrophoresis technology. Differentially expressed proteins were identified using matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and high performance liquid chromatography-electrospray tandem MS (NanoUPLC-ESI-MS/MS). 5 protein spots, for cytokeratin 9, keratin 1, deoxyuridine triphosphatase (dUTPase), aarF domain containing kinase 4 (ADCK 4) and cofilin1, were identified to be significantly changed in COC1/DDP compared with its parental cells. The expression of these five proteins was further validated by quantitative PCR and Western blotting, confirming the results of proteomic analysis. Further research on these proteins may help to identify novel resistant biomarkers or reveal the mechanism of cisplatin-resistance in human ovarian cancers.

• Animal Bioscience
• /
• v.34 no.5
• /
• pp.922-930
• /
• 2021

Objective: Tibetan pigs, predominantly originating from the Tibetan Plateau, have been subjected to long-term natural selection in an extreme environment. To characterize the metabolic adaptations to hypoxic conditions, transcriptomic and proteomic expression patterns in the livers of Tibetan and Yorkshire pigs were compared. Methods: RNA and protein were extracted from liver tissue of Tibetan and Yorkshire pigs (n = 3, each). Differentially expressed genes and proteins were subjected to gene ontology and Kyoto encyclopedia of genes and genomes functional enrichment analyses. Results: In the RNA-Seq and isobaric tags for relative and absolute quantitation analyses, a total of 18,791 genes and 3,390 proteins were detected and compared. Of these, 273 and 257 differentially expressed genes and proteins were identified. Evidence from functional enrichment analysis showed that many genes were involved in metabolic processes. The combined transcriptomic and proteomic analyses revealed that small molecular biosynthesis, metabolic processes, and organic hydroxyl compound metabolic processes were the major processes operating differently in the two breeds. The important genes include retinol dehydrogenase 16, adenine phosphoribosyltransferase, prenylcysteine oxidase 1, sorbin and SH3 domain containing 2, ENSSSCG00000036224, perilipin 2, ladinin 1, kynurenine aminotransferase 1, and dimethylarginine dimethylaminohydrolase 1. Conclusion: The findings of this study provide novel insight into the high-altitude metabolic adaptation of Tibetan pigs.

• Journal of Ginseng Research
• /
• v.43 no.4
• /
• pp.666-675
• /
• 2019

Background: Korean Red Ginseng (KRG) has been widely used as an herbal medicine to normalize and strengthen body functions. Although many researchers have focused on the biological effects of KRG, more studies on the action mechanism of red ginseng are still needed. Previously, we investigated the proteomic changes of the rat spleen while searching for molecular signatures and the action mechanism of KRG. The proteomic analysis revealed that differentially expressed proteins (DEPs) were involved in the increased immune response and phagocytosis. The aim of this study was to evaluate the biological activities of KRG, especially the immune-enhancing response of KRG. Methods: Rats were divided into 4 groups: 0 (control group), 500, 1000, and 2000 mg/kg administration of KRG powder for 6 weeks, respectively. Isobaric tags for relative and absolute quantitation was performed with Q-Exactive LC-MS/MS to compare associated proteins between the groups. The putative DEPs were identified by a current UniProt rat protein database search and by the Gene Ontology annotations. Results: The DEPs appear to increase the innate and acquired immunity as well as immune cell movement. These results suggest that KRG can stimulate immune responses. This analysis refined our targets of interest to include the potential functions of KRG. Furthermore, we validated the potential molecular targets of the functions, representatively LCN2, CRAMP, and HLA-DQB1, by Western blotting. Conclusion: These results may provide molecular signature candidates to elucidate the mechanisms of the immune response by KRG. Here, we demonstrate a strategy of tissue proteomics for the discovery of the molecular function of KRG.

• The Plant Pathology Journal
• /
• v.37 no.1
• /
• pp.36-46
• /
• 2021

Acidovorax citrulli (Ac) is the causal agent of bacterial fruit blotch (BFB) in watermelon, a disease that poses a serious threat to watermelon production. Because of the lack of resistant cultivars against BFB, virulence factors or mechanisms need to be elucidated to control the disease. Glycerol-3-phosphate dehydrogenase is the enzyme involved in glycerol production from glucose during glycolysis. In this study, we report the functions of a putative glycerol-3-phosphate dehydrogenase in Ac (GlpdAc) using comparative proteomic analysis and phenotypic observation. A glpdAc knockout mutant, AcΔglpdAc(EV), lost virulence against watermelon in two pathogenicity tests. The putative 3D structure and amino acid sequence of GlpdAc showed high similarity with glycerol-3-phosphate dehydrogenases from other bacteria. Comparative proteomic analysis revealed that many proteins related to various metabolic pathways, including carbohydrate metabolism, were affected by GlpdAc. Although AcΔglpdAc(EV) could not use glucose as a sole carbon source, it showed growth in the presence of glycerol, indicating that GlpdAc is involved in glycolysis. AcΔglpdAc(EV) also displayed higher cell-to-cell aggregation than the wild-type bacteria, and tolerance to osmotic stress and ciprofloxacin was reduced and enhanced in the mutant, respectively. These results indicate that GlpdAc is involved in glycerol metabolism and other mechanisms, including virulence, demonstrating that the protein has pleiotropic effects. Our study expands the understanding of the functions of proteins associated with virulence in Ac.

• Biotechnology and Bioprocess Engineering:BBE
• /
• v.9 no.2
• /
• pp.69-75
• /
• 2004

In recent years, the importance of proteomic works, such as protein expression, detection and identification, has grown in the fields of proteomic and diagnostic research. This is because complete genome sequences of humans, and other organisms, progress as cellular processing and controlling are performed by proteins as well as DNA or RNA. However, conventional I protein analyses are time-consuming; therefore, high throughput protein analysis methods, which allow fast, direct and quantitative detection, are needed. These are so-called protein microarrays or protein chips, which have been developed to fulfill the need for high-throughput protein analyses. Although protein arrays are still in their infancy, technical development in immobilizing proteins in their native conformation on arrays, and the development of more sensitive detection methods, will facilitate the rapid deployment of protein arrays as high-throughput protein assay tools in proteomics and diagnostics. This review summarizes the basic technologies that are needed in the fabrication of protein arrays and their recent applications.