• Journal of Microbiology and Biotechnology
• /
• v.20 no.2
• /
• pp.350-355
• /
• 2010

Among human antimicrobial peptides (hAMPs), DCD-1L has a broad spectrum of antimicrobial activity over a wide pH range and in high salt concentrations. It offers a promising alternative to conventional antibiotics. The 458-bp-long dermcidin cDNA was amplified by PCR using a human fetal cDNA library as a template. The 147-bp fragment of the MDCD-1L gene encoding an additional methionine residue was subcloned into the pTYB11 vector. Recombinant MDCD-1L was expressed as an intein fusion protein in E. coli, and then purified by affinity chromatography using chitin beads. A small peptide with a molecular mass of about 5 kDa was detected by tricine gel electrophoresis. The recombinant MDCD-1L peptide was purified from the gel and its amino acid sequence was determined by nanoLC-ESI-MS/MS analysis. The initiating amino acid, methionine, remained attached to the N-terminal region of recombinant MDCD-1L. Purified MDCD-1L showed antimicrobial activity against a Micrococcus luteus test strain.

• Archives of Pharmacal Research
• /
• v.24 no.5
• /
• pp.446-454
• /
• 2001

Peptide fragments, isolated from proteolytic cleavage of thyroglobulin at specific sites, were examined for the iodination of tyrosine residues. The 50 kDa polypeptide, which was prepared from digestion of bovine thyroglobulin and continuous preparative SDS-PAGE, was subjected to reduction with DTT and alkylation with iodoacetic acid to generate S-car-boxymethylated peptide derivative, which was further hydrohysed by endoproteinase-Asp-N. Peptide products were separated by RP-HPLC, and each fraction was analyzed by LC/ESI-MS and MALDI-MS analyses. Based on the specificity of endoproteinase-Asp-N andthe mass spectra data, a peptide fragment turned out to correspond to a peptide, DALCCVKCPEGSYFQ (1438-1452), characterized by the presence of a thioredoxin box (CVKC) and a tyrosine residue. In addition, another peptide fragment (1453-1465) containing a thioredoxin box (CIPC) and a tyrosine residue was also observed. However, any evidence of iodination of the tyrosine residue present in these peptides was not provided. Meanwhile, tyrosine residues in the peptides, DVEEALAGKYLAGRFA (1366-1381) and DYSGLLLAFQVFLL (1290-1303) were found to be iodinated; mono- or diiodinated tyrosine residues, characteristic of a hormogenic site, existed in both peptides. In addition, the tyrosine residue in the peptide (1218-1252), corresponding to a hormonogenic site was also iodinated. Thus, there was a sharp difference of the susceptibility to oxidative iodination between the tyrosine residue in a hormonogenic site and that in a thioredoxin region. From these results, it is suggested that polypeptide region adjacent to tyrosine residues may govern the susceptibility of tyrosine to oxidative iodination.

• Asian Pacific Journal of Cancer Prevention
• /
• v.13 no.7
• /
• pp.3265-3270
• /
• 2012

Clinically, elevated cancer antigen 125 (CA-125) in blood predicts tumor burden in a woman's body, especially in the ovary, but cannot differentiate between malignant or benign. We here used intensive modern proteomic approaches to identify predictive proteins in the serum of women with elevated CA-125 to differentiate malignant from benign ovarian tumors. We identified differentially expressed proteins in serum samples of ovarian cancer (OC) patients, benign ovarian tumor (BT) patients, and healthy control women using mass spectrometry-based quantitative proteomics. Both the OC and BT patients had elevated CA-125. Quantitation was achieved using isobaric tags for relative and absolute quantitation. We obtained 124 quantified differential serum proteins in OC compared with BT. Two proteins, apolipoprotein A-4 (APOA4) and natural resistance-associated macrophage 1, were verified using Western blotting. Proteome profiling applied to OC cases identified several differential serum proteins in the serum of women with elevated CA-125. A novel protein, APOA4, has the potential to be a marker for malignant tumor differentiation in the serum of women with elevated CA-125.

• Journal of Microbiology and Biotechnology
• /
• v.21 no.9
• /
• pp.930-936
• /
• 2011

Two new cyclic depsipeptides wewakamide A (1) and guineamide G (2) have been isolated from the marine cyanobacterium Lyngbya semiplena and Lyngbya majuscula, respectively, collected from Papua New Guinea. The amino and hydroxy acid partial structures of wewakamide A and guineamide G were elucidated through extensive spectroscopic techniques, including HR-FABMS, 1D $^1H$ and $^{13}C$ NMR, as well as 2D COSY, HSQC, HSQC-TOCSY, and HMBC spectra. The sequence of the residues of wewakamide A was determined through a combination of ESI-MS/MS, HMBC, and ROESY. Wewakamide A possesses a ${\beta}$-amino acid, 3-amino-2-methylbutanoic acid (Maba) residue, which has only been previously identified in two natural products, guineamide B (3) and dolastatin D (4). Although both new compounds (1,2) showed potent brine shrimp toxicity, only guineamide G displayed significant cytotoxicity to a mouse neuroblastoma cell line with $LC_{50}$ values of 2.7 ${\mu}M$.

• Bulletin of the Korean Chemical Society
• /
• v.25 no.12
• /
• pp.1833-1839
• /
• 2004

Recently mass spectrometry and separation methods such as liquid chromatography have become major tools in the field of proteomics. In this report, we describe in detail our efforts to develop ultra-high pressure capillary reverse-phase liquid chromatography (cRPLC) and its online coupling to a mass spectrometer by a nanoelectrospray (nanoESI) interface. The RPLC system is constructed in house to deliver LC solvents at the pressure up to 20,000 psig, which is four times higher than conventional RPLC systems. The high operation pressure allows the efficient use of packed micro-capillary columns (50, 75 and 150 ${\mu}$m i.d., up to 1.5 m long). We will discuss the effect of column diameter on the sensitivity of cRPLC/MS/MS experiments and the utility of the developed technique for proteome analysis by its application in the analysis of proteome samples having different levels of complexity.

• Korean Journal of Medicinal Crop Science
• /
• v.17 no.6
• /
• pp.464-469
• /
• 2009

This study was conducted to find out environment-friendly disease control method to Alternaria blight caused by Alternaria panax of ginseng. For this study, 150 methanol extracts from medicinal plants were evaluated and the extract of Impatiens balsamina showed most strong antifungal activity against A. panax. The methanol extract of I. balsamina showed also stable antifungal activity against other ginseng pathogens such as Botrytis cinerea and Fusarium sp., when treated by heat or pH. In vivo, Alternaria blight incidence rate was low of 13% with the treatment of I. balsamina methanol extract compared to 35% of the non-treatment. The antifungal compound of I. balsamina was purified and identified by using a silica gel column chromatography, TLC and ESI-LC/MS/MS analysis. The compound which showed strong antifungal activity was identified as 2-methoxy-1,4-naphtoquinone ($C_{11}H_8O_3$).

• Journal of Microbiology
• /
• v.45 no.4
• /
• pp.326-332
• /
• 2007

In order to understand the functional role of CPRl in Saccharomyces cerevisiae KNU5377 with regard to its multi-tolerance characteristics against high temperatures, inorganic acids, and oxidative stress conditions, whole cellular proteins were analyzed via liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). This procedure was followed by two-dimensional (2-D) gel electrophoresis. Under menadione stress conditions, the 23 upregulated proteins were clearly identified only in the wild- type strain of KNU5377. Among the proteins, Sodl1p Tsa1p, Ahp1, Cpr1p, Cpr3, Ssb2p, and Hsp12p were identified as components of antioxidant systems or protein-folding related systems. The CPR1 protein could not be completely detected in the $cpr1{\Delta}$ mutant of KNU5377 and the other upregulated proteins in the wild-type strain evidenced a clear correlation with the results of immunoblot analysis. Moreover, a reduction in growth patterns (about 50%) could be observed in the $cpr1{\Delta}$ mutant, as compared with that of the wild-type strain under mild MD stress conditions. These results indicate that the upregulation of CPR1 may contribute to tolerance against MD as an inducer of oxidative stress.

• Journal of Applied Biological Chemistry
• /
• v.62 no.1
• /
• pp.81-86
• /
• 2019

A yellow resin (gamboge) from Garcinia hanburyi has been widely used as folk medicine due to its antibacterial and antitumor activities. We isolated four ${\alpha}$-glucosidase inhibitory compounds from the methanol extract of gamboge. The compounds (1-4) were identified as gambogoic acid (1), moreollic acid (2), gambogic acid (3), and 10-methoxygambogenic acid (4), respectively through spectroscopic data including 2D-NMR and HREIMS. All compounds were examined in the enzyme inhibition assay against ${\alpha}$-glucosidase to identify their inhibitory potencies and kinetic behavior. All compounds (1-4) showed enzyme inhibition against ${\alpha}$-glucosidase, but the activity was significantly affected by the methoxy group on C-10 of ring A and pentenyl pyran moiety of ring D. For example, compound 1 ($IC_{50}=41.4{\mu}M$) bearing pyran ring eight times effective that 4 ($IC_{50}=350.6{\mu}M$) having geranyl group itself. Most active compound was found out to be gambogoic acid (1) which was analyzed most abundant metabolite in gamboge by LC-ESI-MS/MS. In kinetic study, compounds 1 and 2 were proved as noncompetitive inhibitors.

• Proceedings of the Korean Society of Crop Science Conference
• /
• 2021.04a
• /
• pp.137-137
• /
• 2021

The study of total phenol (TP) content from 700 rice varieties was evaluated using the UV-Vis spectrophotometric method. The calibration curve of serial diluted gallic acid as a standard of this study showed the acceptable performances (R² = 0.999, mean accuracy 90%) and the mean of % relative standard deviation (%RSD: 0.07%) within the range of 7.8 to 1000 ppm concentrations. The mean value of total phenol content from 700 rice varieties was 2723.15 ㎍/g ranged from 55.48 ㎍/g to 9922.23 ㎍/g and the mean %RSD was 2.5%. Furthermore, this study aim was to analyze and profile individual phenolic compounds in the rice genetic resources to construct an integrative database for development of new rice variety with high functionality for health and understanding of phenolics characteristics in the rice grain. Herein, we analyzed selected 100 rice varieties based on high TP content and identified total 15 phenolic compounds by LC-ESI-MS/MS. Among selected 100 rice genetic resources, the phenolic metabolites consisted of higher amount of flavonoid (catechin) and phenolic acid mainly protocatechuic acid. Further research of more selected rice genetic resources would be continued to provide for an integrative phenolics profile of different rice genetic resources.

• BMB Reports
• /
• v.57 no.3
• /
• pp.143-148
• /
• 2024

Pulmonary fibrosis is a serious lung disease that occurs predominantly in men. Genistein is an important natural soybean-derived phytoestrogen that affects various biological functions, such as cell migration and fibrosis. However, the antifibrotic effects of genistein on pulmonary fibrosis are largely unknown. The antifibrotic effects of genistein were evaluated using in vitro and in vivo models of lung fibrosis. Proteomic data were analyzed using nano-LC-ESI-MS/MS. Genistein significantly reduced transforming growth factor (TGF)-β1-induced expression of collagen type I and α-smooth muscle actin (SMA) in MRC-5 cells and primary fibroblasts from patients with idiopathic pulmonary fibrosis (IPF). Genistein also reduced TGF-β1-induced expression of p-Smad2/3 and p-p38 MAPK in fibroblast models. Comprehensive protein analysis confirmed that genistein exerted an anti-fibrotic effect by regulating various molecular mechanisms, such as unfolded protein response, epithelial mesenchymal transition (EMT), mammalian target of rapamycin complex 1 (mTORC1) signaling, cell death, and several metabolic pathways. Genistein was also found to decrease hydroxyproline levels in the lungs of BLM-treated mice. Genistein exerted an anti-fibrotic effect by preventing fibroblast activation, suggesting that genistein could be developed as a pharmacological agent for the prevention and treatment of pulmonary fibrosis.