• Title/Summary/Keyword: MALDI-TOF/TOF MS

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Characterization of a Fibrinolytic Metalloenzyme from a Wild Mushroom, Tricholoma sejunctum (쓴송이버섯으로부터 분리한 혈전용해 금속효소의 특성 연구)

  • Kim, Jun-Ho;Cho, Seung-Koo
    • The Korean Journal of Mycology
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    • v.32 no.2
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    • pp.119-124
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    • 2004
  • Metalloenzyme was purified from the fruiting bodies of Tricholoma sejunctum. MALDI-TOF and ICP/MS analyses revealed that the enzyme had a molecular weight of 18788.25 and includes $Zn^{2+}$ ion. The N-terminal amino acid sequence of the enzyme was Ala-Thr-Tyr-Lys-Ile-X-Ser-Ala-Thr-His-Gln-X-X-Leu-Val. The activity of the enzyme was inhibited by EDTA and 1,10-phenanthroline, indicating that the enzyme was a metalloprotease. No inhibition was found with E-64 and pepstatin. It has broad substrate specificity for synthetic peptides. The enzyme was stable up to $40^{\circ}C$. The activity of the enzyme was increased by $Zn^{2+}$ and $Co^{2+}$, while it was totally inhibited by $Hg^{2+}$. The enzyme hydrolyzes $A{\alpha}$ subunit of human fibrinogen but did not show any reactivity for $B{\beta}$ and ${\gamma}$ form of human fibrinogen.

Characteristics of Wood Tar Produced as Byproduct from Two Types of The Kiln in The Manufacture of Oak Charcoal

  • Yang, Bong Suk;Yang, Jiwook;Kim, Dae-Young;Kim, Jin-Kyu;Hwang, Won-Jung;Kwon, Gu-Joong
    • Journal of the Korean Wood Science and Technology
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    • v.45 no.6
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    • pp.772-786
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    • 2017
  • This study investigated the characteristics of wood tar, produced as a byproduct during the production of charcoal using oak wood by the modified traditional kiln and mechanical steel kiln. The wood tar was analyzed with a number of techniques, including Py-GC/MS, NMR, MALDI-TOF, FT-IR, TG and DSC. The Py-GC/MS analysis indicated that modified traditional kiln generated a higher hydrocarbon ratio in the wood tar than that of mechanical steel kiln. On the other hand, mechanical steel kiln resulted in a higher proportion of phenolic and aromatic hydrocarbon components than that of modified traditional kiln. Those results were also confirmed by NMR analysis. The MALDI-TOF analysis suggested that the wood tar produced in the mechanical steel kiln had a slightly higher molecular weight than the wood tar produced in the modified traditional kiln. In addition, the FT-IR analysis showed characteristic peak of symmetrical stretching vibration of $CH_3$ from the modified traditional kiln while characteristic peaks of the C-C and C-O stretching vibration were observed from the mechanical steel kiln. Moreover, TG and DSC analysis suggested that the mechanical steel kiln is more thermally stable than that of modified traditional kiln. Those findings clearly showed that the method of making charcoal greatly affects the properties of wood tar.

Enzymatic Production of Alkyl β-Glucoside Based on Transglycosylation Activity of Celluclast (효소적 당전이 반응을 이용한 Alkyl β-Glucoside의 생산)

  • Yong, Hwan-Ung;Kim, Seonmi;Shim, Jae-Hoon
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.41 no.10
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    • pp.1417-1422
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    • 2012
  • Alkyl glucosides were synthesized using the transglycosylation reaction of Celluclast, the cellulase from Trichoderma reesei, with cellobiose and various alcohols. Glucose as a by-product of the reaction was removed using the immobilized yeast system. Among the alkyl glucoside products, the acceptor products of methanol and ethanol were confirmed as methyl ${\beta}$-D-glucopyranoside and ethyl ${\beta}$-D-glucopyranoside via MALDI-TOF MS and enzymatic analysis. Optimal yields of methyl ${\beta}$-glucoside and ethyl ${\beta}$-glucoside were 65.3% (mol/mol) and 59.0% (mol/mol), respectively, based on cellobiose consumed.

Comparative proteomics and global genome-wide expression data implicate role of ARMC8 in lung cancer

  • Amin, Asif;Bukhari, Shoiab;Mokhdomi, Taseem A;Anjum, Naveed;Wafai, Asrar H;Wani, Zubair;Manzoor, Saima;Koul, Aabid M;Amin, Basit;Qurat-ul-Ain, Qurat-ul-Ain;Qazi, Hilal;Tyub, Sumira;Lone, Ghulam Nabi;Qadri, Raies A
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.9
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    • pp.3691-3696
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    • 2015
  • Background: Cancer loci comprise heterogeneous cell populations with diverse cellular secretions. Therefore, disseminating cancer-specific or cancer-associated protein antigens from tissue lysates could only be marginally correct, if otherwise not validated against precise standards. Materials and Methods: In this study, 2DE proteomic profiles were examined from lysates of 13 lung-adenocarcinoma tissue samples and matched against the A549 cell line proteome. A549 matched-cancer-specific hits were analyzed and characterized by MALDI-TOF/MS. Results: Comparative analysis identified a total of 13 protein spots with differential expression. These proteins were found to be involved in critical cellular functions regulating pyrimidine metabolism, pentose phosphate pathway and integrin signaling. Gene ontology based analysis classified majority of protein hits responsible for metabolic processes. Among these, only a single non-predictive protein spot was found to be a cancer cell specific hit, identified as Armadillo repeat-containing protein 8 (ARMC8). Pathway reconstruction studies showed that ARMC8 lies at the centre of cancer metabolic pathways. Conclusions: The findings in this report are suggestive of a regulatory role of ARMC8 in control of proliferation and differentiation in lung adenocarcinomas.

Biomarkers Screening Between Preoperative and Postoperative Patients in Pancreatic Cancer

  • Li, Pei;Yang, Juan;Ma, Qing-Yong;Wu, Zheng;Huang, Chen;Li, Xu-Qi;Wang, Zheng
    • Asian Pacific Journal of Cancer Prevention
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    • v.14 no.7
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    • pp.4161-4165
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    • 2013
  • Objective: To investigate discriminating protein patterns and potential biomarkers in serum samples between pre/postoperative pancreatic cancer patients and healthy controls. Methods: 23 serum samples from PC patients (12 preoperative and 11 postoperative) and 76 from healthy controls were analyzed using matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDI-TOF MS) technique combined with magnetic beads-based weak cation-exchange chromatography (MB-WCX). ClinProTools software selected several markers that made a distinction between pancreatic cancer patients and healthy controls. Results: 49 m/z distinctive peaks were found among the three groups, of which 33 significant peaks with a P < 0.001 were detected. Two proteins could distinguish the preoperative pancreatic cancer patients from the healthy controls. About 15 proteins may be potential biomarkers in assessment of pancreatic cancer resection. Conclusion: MB-MALDI-TOF-MS method could generate serum peptidome profiles of pancreatic cancer and provide a new approach to identify potential biomarkers for diagnosis and prognosis of this malignancy.

Proteomic Approach of the Protein Profiles during Seed Maturation in Common Buckwheat (Fagopyrum esculentum Moench.)

  • Park, Min-Hwa;Shin, Dong-Hoon;Han, Myoung-Hae;Yun, Young-Ho;Bae, Jeong-Sook;Lee, Yun-Sang;Chung, Keun-Yook;Lee, Moon-Soon;Woo, Sun-Hee
    • Korean Journal of Plant Resources
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    • v.22 no.3
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    • pp.227-235
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    • 2009
  • Single seeds of common buckwheat cultivar Suwon No. 1 when subjected to SDS-PAGE revealed very high polymorphism. High variation existed for protein or protein subunits with molecular weight 54-47kDa, 45-25kDa and 16-11kDa. The electrophoregram showed variation for globulin as well as other protein fractions. About 300 proteins were separated by two-dimensional electrophoresis in common buckwheat (Fagopyrum esculentum Moench.) seed. Seed maturation is a dynamic and temporally regulated phase of seed development that determines the composition of storage proteins reserves in mature seeds. Buckwheat seeds from 5, 10, 15, 20, and 25 days after pollination and matured stage were used for the analysis. This led to the establishment of high-resolution proteome reference maps, expression profiles of 48 spots. It was identified 48 proteins from MALDI-TOF/MS analysis of wild buckwheat seed storage proteins. The 48 proteins were found identical or similar to those of proteins reported in buckwheat and other plants; it is belonging to 9 major functional categories including seed storage proteins, stress/defense response, protein synthesis, photosynthesis, allergy proteins, amino acid, enzyme, metabolism, and miscellaneous. It appears that the major allergenic storage protein separated played the important role in buckwheat breeding and biochemical characterization.

Determination of Stereochemical Structure of a Grandidentatin Isomer from Populus alba × glandulosa Bark (현사시나무 수피에서 분리한 Grandidentatin Isomer의 입체구조결정)

  • Kwon, Dong-Joo;Kim, Hyun-Seok;Lee, Phil-Ho;Bae, Young-Soo
    • Journal of the Korean Wood Science and Technology
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    • v.37 no.1
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    • pp.114-120
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    • 2009
  • The bark of Populus alba ${\times}$ glandulosa was collected, air-dried and extracted with 70% aqueous acetone. Then it was successively partitioned with n-hexane, $CH_2Cl_2$, EtOAc and $H_2O$. Repeated Sephadex LH-20 column chromatography and preparative TLC on the EtOAc soluble fraction gave a grandidentatin isomer. The structure was elucidated as grandidentatin A (cis-2-hydroxycyclohexyl 6-O-p-coumaroyl-${\beta}$-D-glucopyranoside) on the basis of spectroscopic evidences such as $^1H$-NMR, $^{13}C$-NMR, 2D-NMR and MALDI TOF-MS spectrum followed by acid hydrolysis. Grandidentatin A was identified here for the first time in Populus alba ${\times}$ glandulosa bark, and to the bset of our knowledge it has not been reported in any other literature.

Fibrinolytic Activity and Proteomic Analysis of Bacillus licheniformis HK-12 Isolated from Chungkuk-Jang (청국장에서 분리한 Bacillus licheniformis HK-12의 혈전용해활성과 프로테옴 분석)

  • Sohn, Byung-Hee;Kwon, Sang-Chul;Oh, Kye-Heon
    • Journal of the Korea Academia-Industrial cooperation Society
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    • v.9 no.3
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    • pp.800-806
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    • 2008
  • The strain HK-12 was enriched and isolated from naturally fermented soybean for the production of fibrinolytic enzyme and the proteome of this enzyme induced during the incubation period was analyzed. The activity of fibrinolytic enzyme derived from supernatants of the HK-12 culture was performed by fibrin plate method for solid fibrinolytic activity. As the result, the fibrinolytic activity of HK-12 grown on the nutrient agar media was about 2.3 times greater than that of plasmin used as standard. The purified enzyme was prepared by a series of purification process including ammonium sulfate precipitation, DEAE-cellulose, Sephadex chromatography. The molecular weight of the enzyme was determined to approximately 23kDa with SDS-PAGE. In order to examine which strain HK-12 proteins increased or decreased during the incubation period, 2-DE analysis was performed. Protein spot #1 significantly expressed on the 2-DE gel of bacteria cultivated for 36-hrs was analysed. As the result of protein sequence analysis using MALDI-TOF MS, one protein was identified as serine protein kinase (PrkA).

A Comparison of Genospecies of Clinical Isolates in the Acinetobacter spp. Complex Obtained from Hospitalized Patients in Busan, Korea

  • Park, Gyu-Nam;Kang, Hye-Sook;Kim, Hye-Ran;Jung, Bo-Kyung;Kim, Do-Hee;Chang, Kyung-Soo
    • Biomedical Science Letters
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    • v.25 no.1
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    • pp.40-53
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    • 2019
  • Of the Acinetobacter spp., A. baumannii (genospecies 2) is the most clinically significant in terms of hospital-acquired infections worldwide. It is difficult to perform Acinetobacter-related taxonomy using phenotypic characteristics and routine laboratory methods owing to clusters of closely related species. The ability to accurately identify Acinetobacter spp. is clinically important because antimicrobial susceptibility and clinical relevance differs significantly among the different genospecies. Based on the medical importance of pathogenic Acinetobacter spp., the distribution and characterization of Acinetobacter spp. isolates from 123 clinical samples was determined in the current study using four typically applied bacterial identification methods; partial rpoB gene sequencing, amplified rRNA gene restriction analysis (ARDRA) of the intergenic transcribed spacer (ITS) region of the 16~23S rRNA, the $VITEK^{(R)}$ 2 system (an automated microbial identification system) and matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS). A. baumannii isolates (74.8%, 92/123) were the most common species, A. nosocomialis (10.6%, 13/123) and A. pittii isolates (7.5%, 9/123) were second and third most common strains of the A. calcoaceticus-A. baumannii (ACB) complex, respectively. A. soli (5.0%, 6/123) was the most common species of the non-ACB complex. RpoB gene sequencing and ARDRA of the ITS region were demonstrated to lead to more accurate species identification than the other methods of analysis used in this study. These results suggest that the use of rpoB genotyping and ARDRA of the ITS region is useful for the species-level identification of Acinetobacter isolates.

Studies on Chemical Strutures and Adhesion Performance of pMDI Adhesives Modified by Ozonized Soybean Oil with Different Mixing Ratios (오존산화 콩기름의 구조분석 및 이를 이용한 변성 pMDI 접착제의 중량비에 따른 접착력 변화)

  • You, Young Sam;Lee, Hyun Jong;Lee, Taek Jun;Park, Heon
    • Journal of the Korean Wood Science and Technology
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    • v.37 no.1
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    • pp.56-64
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    • 2009
  • The purpose of this study was to investigate and develop an eco-friendly wood adhesive based on vegetable oil (especially soybean oil), the renewable and sustainable natural resources, using ozonification technology for the chemical structure modification. The soybean oil (SBO) was reacted with $O_3$ at the rate of 7.13 g/h for different times, 15 minutes, 30 minutes, 60 minutes, and 120 minutes. The investigation of the modified chemical structure of the ozonized SBOs were conducted using FT-IR, $^1H$-NMR, MALDI-TOF MS, and GC/MS. As ozonification time increased, the peak of the unsaturated double bonds was disappeared and aldehyde or carboxyl peak appeared because ozonification broke the oil into small molecules. The plywoods were made at $110^{\circ}C$ with 30 seconds/mm hot-press time using the different ozonized SBO/pMDI adhesives and were tested for the dry, wet, cyclic boil test according to the Korea Industrial Standard F3101 Ordinary plywood. The bond strengths gradually increased with increasing ozonification time. The weight ratio 1:1 (ozonized SBO/pMDI), all strengths in 15, 30 and 60 minuets, exceeded constantly the dry, wet, cyclic boiling standard requirement. The range of ozonification time and weight ratio can fulfil1 the requirment of the wet test standard were 30~60 minutes and more than 0.5 pMDI. From the comprehensive view on the results of above experiments, it could be confirmed through experiments that ozonized SBO/pMDI has characteristics of effective reactivity and wet stability showed as an excellent candidate of wood adhesive applications.