• Title/Summary/Keyword: MALDI-TOF/MS

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Characterization of a Blend-Biosurfactant of Glycolipid and Lipopeptide Produced by Bacillus subtilis TU2 Isolated from Underground Oil-Extraction Wastewater

  • Cheng, Fangyu;Tang, Cheng;Yang, Huan;Yu, Huimin;Chen, Yu;Shen, Zhongyao
    • Journal of Microbiology and Biotechnology
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    • v.23 no.3
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    • pp.390-396
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    • 2013
  • Biosurfactants have versatile properties and potential industrial applications. A new producer, B. subtilis TU2, was isolated from the underground oil-extraction wastewater of Shengli Oilfield, China. Preliminary flask culture showed that the titer of biosurfactant obtained from the broth of TU2 was ~1.5 g/l at 48 h (718 mg/l after purification), with a reduced surface tension of 32.5 mN/m. The critical micelle concentration was measured as 50 mg/l and the surface tension maintained stability in solution with 50 g/l NaCl and 16 g/l $CaCl_2$ after 5 days of incubation at $70^{\circ}C$. FT-IR spectra exhibited the structure information of both glycolipid and lipopeptide. MALDI-TOF-MS analyses confirmed that the biosurfactant produced by B. subtilis TU2 was a blend of glycolipid and lipopeptide, including rhamnolipid, surfactin, and fengycin. The blended biosurfactant showed 86% of oil-washing efficiency and fine emulsification activity on crude oil, suggesting its potential application in enhanced oil recovery.

Two-dimensional Electrophoretic Analysis of Nucleotide phosphate Kinase Mediated Hydrogen Peroxide Cross-linking in Saccharamyces cerevisiae (2-D 전기영동 분석을 통한 $H_2O_2$와 연계된 효모 시스템 NDPK에 관한 연구)

  • Moon Hae-Jeong;Yun Dae-Jin;Park Chang-Ho
    • KSBB Journal
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    • v.21 no.1 s.96
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    • pp.16-19
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    • 2006
  • Oxidative modification of nucleoside diphosphate kinase (NDPK) is identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometer. The quaternary structure of NDPK appears to be regulated by cross-linking with an oxidant, $H_2O_2$. We compared roles of NDPK in each of wild type and ynk mutant against oxidative stress. Six specific proteins changed by $H_2O_2$ were identified using two-dimensional electrophoretic analysis. YNK regulated several proteins, related to $H_2O_2$ signaling functions. These results suggest that one of the important functions of NDPK is the regulation of cellular redox state.

Zinc Ions Affect Siderophore Production by Fungi Isolated from the Panax ginseng Rhizosphere

  • Hussein, Khalid Abdallah;Joo, Jin Ho
    • Journal of Microbiology and Biotechnology
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    • v.29 no.1
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    • pp.105-113
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    • 2019
  • Although siderophore compounds are mainly biosynthesized as a response to iron deficiency in the environment, they also bind with other metals. A few studies have been conducted on the impact of heavy metals on the siderophore-mediated iron uptake by microbiome. Here, we investigated siderophore production by a variety of rhizosphere fungi under different concentrations of $Zn^{2+}$ ion. These strains were specifically isolated from the rhizosphere of Panax ginseng (Korean ginseng). The siderophore production of isolated fungi was investigated with chrome azurol S (CAS) assay liquid media amended with different concentrations of $Zn^{2+}$ (50 to $250{\mu}g/ml$). The percentage of siderophore units was quantified using the ultra-violet (UV) irradiation method. The results indicated that high concentrations of $Zn^{2+}$ ion increase the production of siderophore in iron-limited cultures. Maximum siderophore production by the fungal strains was detected at $Zn^{2+}$ ion concentration of $150{\mu}g/ml$ except for Mortierella sp., which had the highest siderophore production at $200{\mu}g/ml$. One potent siderophore-producing strain (Penicillium sp. JJHO) was strongly influenced by the presence of $Zn^{2+}$ ions and showed high identity to P. commune (100% using 18S-rRNA sequencing). The purified siderophores of the Penicillium sp. JJHO strain were chemically identified using UV, Fourier-transform infrared spectroscopy (FTIR), and matrix-assisted laser desorption/ionization time-of-flight mass spectrometer (MALDI-TOF-MS) spectra.

Identification and Antifungal Susceptibility Profiles of Cyberlindnera fabianii in Korea

  • Park, Ji-Hyun;Oh, Junsang;Sang, Hyunkyu;Shrestha, Bhushan;Lee, Hyeyoung;Koo, Jehyun;Cho, Sung-Il;Choi, Ji Seon;Lee, Min-Ha;Kim, Jayoung;Sung, Gi-Ho
    • Mycobiology
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    • v.47 no.4
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    • pp.449-456
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    • 2019
  • Invasive fungal infections caused by Cyberlindnera fabianii have recently increased. However, biochemical kits such as API 20 C AUX and Vitek-2C have misidentified this species as other Candida spp. such as C. pelliculosa or C. utilis due to no information of Cy. fabianii in yeast database. During our 2016-2017 surveys, eleven isolates of Cy. fabianii were obtained in International St. Mary's Hospital in Korea. Here, we describe its morphological and molecular characteristics and tested its antifungal susceptibility against nine antifungal agents. The sequences of the ITS region and the D1/D2 region of LSU revealed 100% identity with the sequences of Cy. fabianii. In comparison with the results from MALDI-TOF mass spectrometry, we found that Cy. fabianii can be distinguished from other species. In antifungal susceptibility test, voriconazole and echinocandins exhibited good antifungal activities against the majority of Cy. fabianii isolates despite the absence of standard criteria.

Purification of Two Novel Antimicrobial Peptides from Pyloric Caeca of the Starfish Asterina pectinifera (별불가사리 Asterina pectinifera의 유문맹낭 추출물로부터 새로운 2종류의 항균활성 펩타이드의 정제)

  • Go, Hye-Jin;Bae, Yun Jung;Park, Nam Gyu
    • Journal of Life Science
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    • v.24 no.8
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    • pp.860-864
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    • 2014
  • PAP-1, a novel antimicrobial peptide isolated from pyloric caeca extract of the starfish Asterina pectinifera was purified and characterized. First, the acidified pyloric caeca extract was put through Sep-Pak C18 solid phase extraction cartridge using a stepwise gradient. Among the eluents, RM 60 (retained materials at 60% methanol) showed good antimicrobial activity against Bacillus subtilis and Escherichia coli D31 and was purified in C18 reversed-phase and ion-exchange high-performance liquid chromatography columns. The purification steps yielded two novel peptides showing strong antimicrobial activities. These peptides were named pyloric caeca A. pectinifera peptide 1 and 2 (PAP-1 and PAP-2). For the characterization of the purified peptides, the molecular weights and amino acid sequences were determined by MALDI-TOF MS and Edman degradation. The molecular weights of PAP-1 and PAP-2 were about 2951.54 Da and 2980.15 Da respectively. The amino acid sequences of PAP-1 and PAP-2 were partially determined: AIQNAGES and AIQNAAES, respectively. PAP-2 is an isoform of PAP-1, differing merely by a single residue at position 6 (glycine or alanine). The comparison of the N-terminal amino acid sequences and molecular weights of the peptides with those of other known antimicrobial peptides revealed that PAP-1 and PAP-2 have no homology with any known peptides. These findings suggest that PAP-1 and PAP-2 play a significant role in the innate defense system of starfish pyloric caeca.

Optimal Conditions and Substrate Specificity for Trehalose Production by Resting Cells of Arthrobacter crystallopoietes N-08

  • Seo, Yi-Seul;Shin, Kwang-Soon
    • Preventive Nutrition and Food Science
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    • v.16 no.4
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    • pp.357-363
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    • 2011
  • Recently, we found that Arthrobacter crystallopoietes N-08 isolated from soil directly produces trehalose from maltose by a resting cell reaction. In this study, the optimal set of conditions and substrate specificity for the trehalose production using resting cells was investigated. Optimum temperature and pH of the resting cell reaction were $55^{\circ}C$ and pH 5.5, respectively, and the reaction was stable for two hours at $37{\sim}55^{\circ}C$ and for one hour at the wide pH ranges of 3~9. Various disaccharide substrates with different glycosidic linkages, such as maltose, isomaltose, cellobiose, nigerose, sophorose, and laminaribiose, were converted into trehalose-like spots in thin layer chromatography (TLC). These results indicated broad substrate specificity of this reaction and the possibility that cellobiose could be converted into other trehalose anomers such as ${\alpha},{\beta}$- and ${\beta},{\beta}$-trehalose. Therefore, the product after the resting cell reaction with cellobiose was purified by ${\beta}$-glucosidase treatment and Dowex-1 ($OH^-$) column chromatography and its structure was analyzed. Component sugar and methylation analyses indicated that this cellobiose-conversion product was composed of only non-reducing terminal glucopyranoside. MALDI-TOF and ESI-MS/MS analyses suggested that this oligosaccharide contained a non-reducing disaccharide unit with a 1,1-glucosidic linkage. When this disaccharide was analyzed by $^1H$-NMR and $^{13}C$-NMR, it gave the same signals with ${\alpha}$-D-glucopyranosyl-(1,1)-${\alpha}$-D-glucopyranoside. These results suggest that cellobiose can be converted to ${\alpha},{\alpha}$-trehalose by the resting cells of A. crystallopoietes N-08.

Proteomic Analysis of Protein Expression in Streptococcus pneumoniae in Response to Temperature Shift

  • Lee Myoung-Ro;Bae Song-Mee;Kim Tong-Soo;Lee Kwang-Jun
    • Journal of Microbiology
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    • v.44 no.4
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    • pp.375-382
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    • 2006
  • From its initial colonization to causation of disease, Streptococcus pneumoniae has evolved strategies to cope with a number of stressful in vivo environmental conditions. In order to analyze a global view of this organism's response to heat shock, we established a 2-D electrophoresis proteome map of the S. pneumoniae D39 soluble proteins under in vitro culture conditions and performed the comparative proteome analysis to a 37 to $42^{\circ}C$ temperature up-shift in S. pneumoniae. When the temperature of an exponentially growing S. pneumoniae D39 culture was raised to $42^{\circ}C$, the expression level of 25 proteins showed changes when compared to the control. Among these 25 proteins, 12 were identified by MALDI-TOF and LC-coupled ESI MS/MS. The identified proteins were shown to be involved in the general stress response, energy metabolism, nucleotide biosynthesis pathways, and purine metabolism. These results provide clues for understanding the mechanism of adaptation to heat shock by S. pneumoniae and may facilitate the assessment of a possible role for these proteins in the physiology and pathogenesis of this pathogen.

Proteomic Analysis of the Triglyceride-Rich Lipoprotein-Laden Foam Cells

  • Lu, Yanjun;Guo, Jianli;Di, Yong;Zong, Yiqiang;Qu, Shen;Tian, Jun
    • Molecules and Cells
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    • v.28 no.3
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    • pp.175-181
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    • 2009
  • In hypertriglyceridaemic individuals, atherosclerogenesis is associated with the increased concentrations of very low density lipoprotein (VLDL) and VLDL-associated remnant particles. In vitro studies have suggested that VLDL induces foam cells formation. To reveal the changes of the proteins expression in the process of foam cells formation induced by VLDL, we performed a proteomic analysis of the foam cells based on the stimulation of differentiated THP-1 cells with VLDL. Using two-dimensional gel electrophoresis (2-DE) and matrix-assisted laser-desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis, 14 differentially expressed proteins, containing 8 up-regulated proteins and 6 down-regulated proteins were identified. The proteins are involved in energy metabolism, oxidative stress, cell growth, differentiation and apoptosis, such as adipose differentiation-related protein (ADRP), enolase, S100A11, heat shock protein 27 and so on. In addition, the expression of some selected proteins was confirmed by Western blot and RT-PCR analysis. The results suggest that VLDL not only induces lipid accumulation, but also brings about foam cells diverse characteristics by altering the expression of various proteins.

Proteomic Analysis of Serum Samples from Natural Herb and Immunoglobulin Yolksac (Ig Y) treated Porcine

  • Park, Hyeon-Soo;Nagappan, Arulkumar;Hong, Gyeong-Eun;Yumnam, Silvia;Lee, Ho-Jung;Kim, Gon-Sup
    • Journal of Korean Clinical Health Science
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    • v.1 no.1
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    • pp.83-89
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    • 2013
  • Purpose. Natural herb has been used for traditional medicine for thousands of years. For this reason, alternative medicine methods affect to domestic animal field. The main purpose of this study was to confirm the regulated proteins after feed additive treatment. Methods. We used total thirty male pigs were used for this experiment. E. coli and Salmonella typhimurium, were used for promote enteritis. Animals were divided into a negative control group, positive control group and test group to determine the effect of an additive mixture on the changes in protein expression in serum. The pigs were given a food supplemented with a natural herbal additive containing immunoglobulin yolksac (IgY) at concentrations 1%. On the 1st week and after eight weeks of feeding, the serums were collected from each group. The changes in the serum proteome as a response to the herbal additive were examined using two-dimensional polyacrylamide gel electrophoresis. Results. A total of 14 differentially expressed protein spots were identified by comparing the protein profiles of the control and additive treated porcine serums. Finally, 7 proteins were detected by MALDI-TOF/MS. Moreover, the proteins detected are involved in a cholesterol control factor, inflammation regulator, Conclusion. These results support of the hypothesis that a natural herbal additive containing IgY can affect the immune regulation system and reduce the pathological process by microbial infections.

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