• Title, Summary, Keyword: fibrinogen alpha chain

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Proteomic Profiling of Serum from Stage I Lung Squamous Cell Carcinoma Patients

  • Li, Xin-Ju;Wu, Qi-Fei;He, Da-Lin;Fu, Jun-Ke;Jin, Xin
    • Asian Pacific Journal of Cancer Prevention
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    • v.14 no.4
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    • pp.2273-2276
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    • 2013
  • Objectives: This study employed proteomic profiling to identify specific tumor markers that might improve early diagnosis of lung squamous cell carcinoma. Methods: Serum samples were isolated from 30 patients with stage I lung squamous cell carcinoma and 30 age-and gender-matched healthy controls, and proteomic profiles were obtained by matrix-assisted laser desorption ionization time of flight mass spectrometry. Results: Three highly expressed potential tumor markers were identified in the sera of stage I lung squamous cell carcinoma patients, with molecular weights of 3261.69, 3192.07, and 2556.92 Da. One protein peak with molecular weight 3261.69 Da was chosen as the candidate biomarker and identified as a fibrinogen alpha chain through a search of the IPI, NCBI or SWISS-PROT protein databases. Conclusion: As a potential tumor biomarker, fibrinogen alpha chain may be applicable for the early diagnosis and prognosis of lung squamous cell carcinoma patients.

Association between Single Nucleotide Polymorphisms of the Fibrinogen Alpha Chain (FGA) Gene and Type 2 Diabetes Mellitus in the Korean Population

  • Hwang, Joo-Yeon;Ryu, Min-Hyung;Go, Min-Jin;Oh, Berm-Seok;Cho, Yoon-Shin
    • Genomics & Informatics
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    • v.7 no.2
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    • pp.57-64
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    • 2009
  • Fibrinogen alpha chain (FGA), a subunit of fibrinogen, might be a potential player for type 2 diabetes mellitus (T2DM), since the plasma levels of fibrinogen is known to be related to the incidence of T2DM. To elucidate the potential role of FGA in T2DM, we investigated whether FGA genetic variations are relevant in T2DM in the Korean population. Seven FGA single nucleotide polymorphisms (SNPs) were genotyped in Ansung and Ansan cohorts (474 T2DM subjects and 470 normal controls) in Korea. The association between SNPs and T2DM was determined by logistic regression analysis. Genetic relevance of SNPs to T2DM-related phenotypes was investigated by multiple linear regression analysis. Statistical analysis revealed that among seven FGA SNPs, significant associations with T2DM were observed in FGA rs2070011 (p=0.013-0.034, OR=0.72${\sim}$0.79), rs6050 (p=0.026${\sim}$0.048, OR=1.24${\sim}$1.37), and rs2070022 (p=0.016${\sim}$0.039, OR=0.70${\sim}$0.72). Two SNPs, rs2070011 and rs6050, also showed significant association with T2DM-related phenotypes such as triglyceride (p=0.005${\sim}$0.011 for rs2070011 and p=0.003${\sim}$0.008 for rs6050), total cholesterol (p=0.01 for rs2070011 and p=0.024 for rs6050) and fasting glucose (p=0.035${\sim}$0.036 for rs2070011 and p=0.048 for rs6050) in 470 normal controls. Our association study implies that FGA might be an important genetic factor in T2DM pathogenesis in the Korean population by affecting plasma lipid and glucose levels.

Characterization of Fibrinolytic Proteases from Gloydius blomhoffii siniticus Venom

  • Choi, Suk-Ho
    • Journal of Pharmacopuncture
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    • v.14 no.3
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    • pp.71-79
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    • 2011
  • Objectives : This study was undertaken to identify fibrinolytic proteases from Gloydius blomhoffii siniticus venom and to characterize a major fibrinolytic protease purified from the venom. Methods : The venom was subjected to chromatography using columns of Q-Sepharose and Sephadex G-75. The molecular weights of fibrinolytic proteases showing fibrinolytic zone in fibrin plate assay were determined in SDS-PAGE (Sodium dodecyl sulfate-polyacrylamide gel electrophoresis) The effects of inhibitors and metal ions on fibrinolytic protease and the proteolysis patterns of fibrinogen, gelatin, and bovine serum albumin were investigated. Results : 1) The fibrinolytic fractions of the three peaks isolated from Gloydius blomhoffii siniticus venom contained two polypeptides of 46 and 59 kDa and three polypeptides of 32, 18, and 15 kDa and a major polypeptide of 54 kDa, respectively. 2) The fibrinolytic activity of the purified protease of 54 kDA was inhibited by metal chelators, such as EDTA, EGTA, and 1,10-phenanthroline, and disulfhydryl-reducing compounds, such as dithiothreitol and cysteine. 3) Calcium chloride promoted the fibrinolytic activity of the protease, but mercuric chloride and cobalt(II) chloride inhibited it. 4) The fibrinolytic protease cleaved preferentially A${\alpha}$-chain and slowly B${\beta}$-chain of fibrinogen. It also hydrolyzed gelatin but not bovine serum albumin. Conclusions : The Gloydius blomhoffii siniticus venom contained more than three fibrinolytic proteases. The major fibrinolytic protease was a metalloprotease which hydrolyzed both fibrinogen and gelatin, but not bovine serum albumin.

Physiological Activities of Water Extract and Solvent Fractions of an Edible Mushroom, Pholiota adiposa (검은비늘버섯 물 추출물 및 유기용매 분획물의 생리활성 효과)

  • Kim, Jun-Ho
    • The Korean Journal of Mycology
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    • v.42 no.3
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    • pp.207-212
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    • 2014
  • This study was conducted in order to investigate the physiological activities, including antioxidative, fibrinolytic, thrombin inhibitory, and ${\alpha}$-glucosidase inhibitory activities of the water extract and solvent fractions isolated from Pholiota adiposa. The antioxidative activities of the water extract and water fraction were 57.57% and 48.27%, respectively. The fibrinolytic activity was strong only in the ethyl acetate fraction at 0.70 plasmin units/mL. The ethyl acetate fraction showed high thrombin inhibitory activity, and a-glucosidase inhibitory activity at 77.67% and 89.32%, respectively. The ethyl acetate fraction hydrolyzed both $A{\alpha}$ and $B{\beta}$ subunits of human fibrinogen, but did not show reactivity for the ${\gamma}$ form of human fibrinogen. Fibrinolytic activity of the ethyl acetate fraction was not decreased by heating for 10 min at $100^{\circ}C$.

Purification and Characterization of Fibrinolytic Enzyme from Tricholoma saponaceum (II) (할미송이버섯으로부터 혈전용해효소의 정제 및 특성 연구 (II))

  • 김준호
    • Biomedical Science Letters
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    • v.6 no.4
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    • pp.261-268
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    • 2000
  • Fibrinolytic enzyme (FE-2) was purified from the fruiting bodies of Tricholoma saponaceum using DEAE-Cellulose chromatography and Mono-S column chromatography, The enzyme has a molecular weight of 18.23 kDa and include Zn$^{2+}$ ion as found by ICP/MS. The N-terminal amino acid sequence of the enzyme was A-L-Y-V-G-X-S-P-X-Q-Q-S-L-L-V It has a pH optimum at pH 7.5, suggested that FE-2 was a neutral pretense. The activity of FE-2 was highly inhibited by EDTA and 1,10-phenanthroline, indicating that the enzyme is a metalloprotease. The activity of FE-2 was increased by $Mg^{2+}$, Zn$^{2+}$, Fe$^{2+}$, and Co$^{2+}$, but the enzyme activity was totally inhibited by Hg$^{2+}$. No inhibition was found with PMSF, E-64, pepstatin and 2-mercaptoethanol. The enzyme hydrolyzed both $A\alpha$ and B$\beta$ chains of human fibrinogen. The $\gamma$ chain was resistant to hydrolysis by FE-2.

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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.

ADHESIVENESS EVALUATION OF ACTIVATED PLATELET USING Arg-Gly-Asp-Phe(RGDF)-IMMOBILIZED SURFACE

  • Kim, J.H.;Kim, H.J.;Kim, J.;Ryu, G.H.;Min, B.G.;Choe, T.B.
    • Proceedings of the KOSOMBE Conference
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    • v.1997 no.11
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    • pp.333-336
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    • 1997
  • The adhesion of activated and normal platelets to fibrinogen requires the receptor binding site of GPIIb/IIIa. These recognition sites exists in the A ${\alpha}$ chain(RGDS at 572-575 and RGDF at 95-98) and the carboxy-terminal of ${\gamma}$ chain (HHLGGAKQAGDV at 400-411) of fibrinogen. In this study, we developed RGDF-immobilized surface to detect the unctional state of platelet. RGDF-immobilized surface was prepared on the glass using photolithographic technology. Platelet adhesion to RGDF-immobilized surface was observed by staining platelets with mepacrine using a fluorescence microscope using mepacrine. Using the RGDF peptide of fragment E, we observed that the platelets pretreated with PGE1 interacted incompletely with RGDF-immobilized surface, whereas ADP activated platelets interacted with the surface extensively. These results show that the distinct selectivity of RGDF-immobilized micro-patterned surface can be used to detect the unctional state of platelets.

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Purification and Characterization of a Serine Protease (CPM-2) with Fibrinolytic Activity from the Dung Beetles

  • Ahn, Mi-Young;Hahn, Bum-Soo;Ryu, Kang-Sun;Hwang, Jae-Sam;Kim, Yeong-Shik
    • Archives of Pharmacal Research
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    • v.28 no.7
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    • pp.816-822
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    • 2005
  • Catharsius protease-2 (CPM-2) was isolated from the body of dung beetles, Catharsius molossus, using a three step purification process (ammonium sulfate fractionation, gel filtration on Bio-Gel P-60, and affinity chromatography on DEAE Affi-Gel blue). The purified CPM-2, having a molecular weight of 24 kDa, was assessed homogeneously by SDS-polyacrylamide gel electrophoresis. The N-terminal amino acid sequence of CPM-2 was composed of X Val Gin Asp Phe Val Glu Glu lie Leu. CPM-2 was inactivated by $Cu^{2+}\;and\;Zn^{2+}$ and strongly inhibited by typical serine proteinase inhibitors such as TLCK, soybean trypsin inhibitor, aprotinin, benzamidine, and ${\alpha}_1$-antitrypsin. However, EDTA, EGTA, cysteine, $\beta$-mercaptoethanol, E64, and elastatinal had little effect on enzyme activity. In addition, antiplasmin and antithrombin III were not sensitive to CPM-2. Based on the results of a fibrinolytic activity test, CPM-2 readily cleaved $A{\alpha}-$ and $B{\beta}$-chains of fibrinogen and fibrin, and y-chain of fibrinogen more slowly. The nonspecific action of the enzyme resulted in extensive hydrolysis, releasing a variety of fibrinopeptides of fibrinogen and fibrin. Polyclonal antibodies of CPM-2 were reactive to the native form of antigen. The ELISA was applied to detect quantities, in nanograms, of the antigen in CPM-2 protein.

Purification and Characterization of a Subtilisin D5, a Fibrinolytic Enzyme of Bacillus amyloliquefaciens DJ-5 Isolated from Doenjang

  • Choi, Nack-Shick;Chung, Dong-Min;Han, Yun-Jon;Kim, Seung-Ho;Song, Jae-Jun
    • Food Science and Biotechnology
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    • v.18 no.2
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    • pp.500-505
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    • 2009
  • The fibrinolytic enzyme, subtilisin D5, was purified from the culture supernatant of the isolated Bacillus amyloliquefaciens DJ-5. The molecular weight of subtilisin D5 was estimated to be 30 kDa. Subtilisin D5 was optimally active at pH 10.0 and $45^{\circ}C$. Subtilisin D5 had high degrading activity for the A$\alpha$-chain of human fibrinogen and hydrolyzed the $B{\beta}$-chain slowly, but did not affect the $\gamma$-chain, indicating that it is an $\alpha$-fibrinogenase. Subtilisin D5 was completely inhibited by phenylmethylsulfonyl fluoride, indicating that it belongs to the serine protease. The specific activity (F/C, fibrinolytic/caseinolytic activity) of subtilisin D5 was 2.37 and 3.52 times higher than those of subtilisin BPN' and Carlsberg, respectively. Subtilisin D5 exhibited high specificity for Meo-Suc-Arg-Pro-Tyr-pNA (S-2586), a synthetic chromogenic substrate for chymotrypsin. The first 15 amino acid residues of the N-terminal sequence of subtilisin D5 are AQSVPYGISQIKAPA; this sequence is identical to that of subtilisin NAT and subtilisin E.

Isolation from Gloydius blomhoffii siniticus Venom of a Fibrin(ogen)olytic Enzyme Consisting of Two Heterogenous Polypeptides

  • Choi, Suk-Ho;Lee, Seung-Bae
    • Journal of Pharmacopuncture
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    • v.16 no.2
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    • pp.46-54
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    • 2013
  • Objective: This study was undertaken to isolate a fibrin(ogen)olytic enzyme from the snake venom of Gloydius blomhoffii siniticus and to investigate the enzymatic characteristics and hemorrhagic activity of the isolated enzyme as a potential pharmacopuncture agent. Methods: The fibrinolytic enzyme was isolated by using chromatography, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and fibrin plate assay. The characteristics of the enzyme were determined by using fibrin plate assay, protein hydrolysis analysis, and hemorrhage assay. Its amino acid composition was determined. Results: The fibrin(ogen)olytic enzyme with the molecular weight of 27 kDa (FE-27kDa) isolated from G. b. siniticus venom consisted of two heterogenous disulfide bond-linked polypeptides with the molecular weights of 15 kDa and 18 kDa. When more than $20{\mu}g$ of FE-27kDa was applied on the fibrin plate, fibrinolysis zone was formed as indicating its fibrinolytic activity. The fibrinolytic activity was inhibited completely by phenylmethanesulfonylfluoride (PMSF) and ethylenediaminetetraacetic acid (EDTA) and partially by thiothreitol and cysteine. Metal ions such as $Hg^{2+}$ and $Fe^{2+}$ inhibited the fibrinolytic activity completely, but $Mn^{2+}$ did not. FE-27kDa preferentially hydrolyzed ${\alpha}$-chain of fibrinogen and slowly hydrolyzed ${\beta}$-chain, but did not hydrolyze ${\gamma}$-chain. High-molecular-weight polypeptides of gelatin were hydrolyzed partially into polypeptides with molecular weights of more than 45 kDa. A dosage of more than $10{\mu}g$ of FE-27kDa per mouse was required to induce hemorrhage beneath the skin. Conclusion: FE-27kDa was a serine proteinase consisting of two heterogeneous polypeptides, hydrolyzed fibrin, fibrinogen, and gelatin, and caused hemorrhage beneath the skin of mouse. This study suggests that the potential of FE-27kDa as pharmacopuncture agent should be limited due to low fibrinolytic activity and a possible side effect of hemorrhage.