• Journal of The Korean Society of Grassland and Forage Science
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• v.34 no.4
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• pp.262-268
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• 2014

In order to reveal the aluminum (Al) stress tolerance mechanisms in alfalfa plant at low pH soil, a proteomic approach has been conducted. Alfalfa plants were exposed to Al stress for 5 days. The plant growth and total chlorophyll content are greatly affected by Al stress. The malondialdehyde (MDA) and $H_2O_2$ contents were increased in a low amount but free proline and soluble sugar contents, and the DPPH-radical scavenging activity were highly increased. These results indicate that antioxidant activity (DPPH activity) and osmoprotectants (proline and sugar) may involve in ROS ($H_2O_2$) homeostasis under Al stress. In proteomic analysis, over 500 protein spots were detected by 2-dimentional gel electrophoresis analysis. Total 17 Al stress-induced proteins were identified, of which 8 protein spots were up-regulated and 9 were down-regulated. The differential expression patterns of protein spots were selected and analyzed by the peptide mass fingerprinting (PMF) using MALDI-TOF MS analysis. Three protein spots corresponding to Rubisco were significantly down-regulated whereas peroxiredoxin and glutamine synthetase were up-regulated in response to Al stress. The different regulation patterns of identified proteins were involved in energy metabolism and antioxidant / ROS detoxification during Al stress in alfalfa. Taken together, these results provide new insight to understand the molecular mechanisms of alfalfa plant in terms of Al stress tolerance.

• Journal of Microbiology and Biotechnology
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• v.18 no.8
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• pp.1368-1376
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• 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 Computing Science and Engineering
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• v.1 no.1
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• pp.1-30
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• 2007

In this paper we have provided an extensive survey of the databases and other resources related to the current research in bioinformatics and the issues that confront the database researcher in helping the biologists. Initially we give an overview of the concepts and principles that are fundamental in understanding the basis of the data that has been captured in these databases. We briefly trace the evolution of biological advances and point out the importance of capturing data about genes, the fundamental building blocks that encode the characteristics of life and proteins that are the essential ingredients for sustaining life. The study of genes and proteins is becoming extremely important and is being known as genomics and proteomics, respectively. Whereas there are numerous databases related to various subfields of biology, we have maintained a focus on genomic and proteomic databases which are the crucial stepping stones for other fields and are expected to play an important role in the future applications of biology and medicine. A detailed listing of these databases with information about their sizes, formats and current status is presented. Related databases like molecular pathways and interconnection network databases are mentioned, but their full coverage would be beyond the scope of a single paper. We comment on the peculiar nature of the data in biology that presents special problems in organizing and accessing these databases. We also discuss the capabilities needed for database development and information management in the bioinformatics arena with particular attention to ontology development. Two research case studies based on our own research are summarized dealing with the development of a new genome database called Mitomap and the creation of a framework for discovery of relationships among genes from the biomedical literature. The paper concludes with an overview of the applications that will be driven from these databases in medicine and healthcare. A glossary of important terms is provided at the end of the paper.

• Journal of Microbiology and Biotechnology
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• v.29 no.11
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• pp.1817-1829
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• 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
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• v.13 no.12
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• pp.6435-6439
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• 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.

• Bulletin of the Korean Chemical Society
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• v.32 no.2
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• pp.599-604
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• 2011

In typical proteomic analysis, trypsin digestion is one of the most time-consuming steps. Conventional proteomic sample preparation methods use an overnight trypsin digestion method. In this study, we compared high-pressure cycling technology (PCT) during enzyme digestion for proteome analysis to the conventional method. We examined the effect of PCT on enzyme activity at temperatures of 25, 37, and $50^{\circ}C$. Although a fast digestion (1 h) was used for the standard protein mixture analysis, the PCT-assisted method with urea showed better results for protein sequence coverage and the number of peptides identified compared with the conventional method. There was no significant difference between temperatures for PCT-assisted digestion; however, we selected PCT-assisted digestion with urea at $25^{\circ}C$ as an optimized method for fast enzyme digestion, based on peptide carbamylation at these conditions. The optimized method was used for stem cell proteome analysis. We identified 233, 264 and 137 proteins using the conventional method with urea at $37^{\circ}C$ for 16h, the PCT-assisted digestion with urea at $25^{\circ}C$ for 1 h, and the non-PCT-assisted digestion with urea at $25^{\circ}C$ for 1 h, respectively. A comparison of these results suggests that PCT enhanced the enzyme digestion by permitting better access to cleavage sites on the proteins.

• Journal of Plant Biotechnology
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• v.47 no.3
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• pp.209-217
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• 2020

The presence of high amounts of seed storage proteins (SSPs) improves the overall quality of soybean seeds. However, these SSPs pose a major limitation due to their high abundance in soybean seeds. Although various technical advancements including mass-spectrometry and bioinformatics resources were reported, only limited information has been derived to date on soybean seeds at proteome level. Here, we applied a tandem mass tags (TMT)-based quantitative proteomic analysis to identify the significantly modulated proteins in the seeds of two soybean cultivars showing varying protein contents. This approach led to the identification of 5,678 proteins of which 13 and 1,133 proteins showed significant changes in Daewon (low-protein content cultivar) and Saedanbaek (high-protein content cultivar) respectively. Functional annotation revealed that proteins with increased abundance in Saedanbaek were mainly associated with the amino acid and protein metabolism involved in protein synthesis, folding, targeting, and degradation. Taken together, the results presented here provide a pipeline for soybean seed proteome analysis and contribute a better understanding of proteomic changes that may lead to alteration in the protein contents in soybean seeds.

• Asian-Australasian Journal of Animal Sciences
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• v.24 no.2
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• pp.272-278
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• 2011

Bovine whey protein expression patterns of colostrum are much different from that of milk. Moreover, bovine colostrum is an important source of protective, nutritional and developmental factors for the newborn. However, to our knowledge, no research has been performed to date using a comparative proteomic method on the changes in the bovine whey proteome during the transition from colostrum to milk. This study therefore separated whey protein of days 1, 3, 7 and 21 after calving using two dimension electrophoresis. Differentially expressed proteins at different collection times were identified using high-performance liquid chromatography in tandem with mass spectrometry (LC/MS) and validated by enzyme-linked immunosorbent assay (ELISA) in order to understand the developmental changes in the bovine whey proteome during the transition from colostrum to milk. The expression patterns of whey protein of days 1 and 3 post-partum were similar except that immunoglobulin G was down-regulated on day 3, and four proteins were found to be down-regulated on days 7 and 21 compared with day 1 after delivering, including immunoglobulin G, immunoglobulin M, albumin, and lactotransferrin, which are involved in immunity and molecule transport. The results of this study confirm the comparative proteomic method has the advantage over other methods such as ELISA and immunoassays in that it can simultaneously detect more differentially expressed proteins. In addition, the difference in composition of milk indicates a need for adjustment of the colostrum feeding regimen to ensure a protective immunological status for newborn calves.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.10
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• pp.6069-6075
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• 2013

Background: At present, the diagnosis of colorectal cancer (CRC) requires a colorectal biopsy which is an invasive procedure. We undertook this pilot study to develop an alternative method and potential new biomarkers for diagnosis, and validated a set of well-integrated tools called ClinProt to investigate the serum peptidome in CRC patients. Methods: Fasting blood samples from 67 patients diagnosed with CRC by histological diagnosis, 55 patients diagnosed with colorectal adenoma by biopsy, and 65 healthy volunteers were collected. Division was into a model construction group and an external validation group randomly. The present work focused on serum proteomic analysis of model construction group by ClinProt Kit combined with mass spectrometry. This approach allowed construction of a peptide pattern able to differentiate the studied populations. An external validation group was used to verify the diagnostic capability of the peptidome pattern blindly. An immunoassay method was used to determine serum CEA of CRC and controls. Results: The results showed 59 differential peptide peaks in CRC, colorectal adenoma and health volunteers. A genetic algorithm was used to set up the classification models. Four of the identified peaks at m/z 797, 810, 4078 and 5343 were used to construct peptidome patterns, achieving an accuracy of 100% (> CEA, P<0.05). Furthermore, the peptidome patterns could differentiate the validation group with high accuracy close to 100%. Conclusions: Our results showed that proteomic analysis of serum with MALDI-TOF MS is a fast and reproducible approach, which may provide a novel approach to screening for CRC.

• Journal of Microbiology and Biotechnology
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• v.24 no.1
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• pp.119-126
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• 2014

Rheumatoid arthritis (RA) is a chronic and systemic inflammatory disorder that primarily affects the flexible joints and may also affect a number of tissues and organs. The progression of RA involves an inflammatory response of the capsule around the joint, swelling of synovial cells with excess synovial fluid (SF), and the development of fibrous tissue in the synovium. Since the progressive pathology of the disease often leads to the irreversible destruction of articular cartilage and ankylosis of the joint, early diagnosis of RA is essential. Thus, we undertook a comparative proteomic approach to investigate novel biomarkers for early diagnosis using SFs and serums from RA patients. As a result, we identified 32 differentially expressed spots in SFs and 34 spots in serums. The differential expression of the STEAP4 and ZNF 658 proteins were validated using immunoblotting of the SFs and serums, respectively. These data suggest that differentially expressed proteins in SFs and serums could be used as RA-specific biomarkers for the diagnosis and monitoring of RA. Furthermore, these findings advance our understanding of the molecular etiopathogenesis of RA.