Navegando por Palavras-chave "thrombin"
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- ItemSomente MetadadadosActivation profiles of human kallikrein-related peptidases by proteases of the thrombostasis axis(Cold Spring Harbor Lab Press, Publications Dept, 2008-11-01) Yoon, Hyesook; Blaber, Sachiko I.; Evans, D. Michael; Trim, Julie; Juliano, Maria Aparecida [UNIFESP]; Scarisbrick, Isobel A.; Blaber, Michael; Florida State Univ; Vantia Ltd; Ferring Res Ltd; Universidade Federal de São Paulo (UNIFESP); Mayo Med & Grad SchThe human kallikrein-related peptidases (KLKs) comprise 15 members (KLK1-15) and are the single largest family of serine proteases. the KLKs are utilized, or proposed, as clinically important biomarkers and therapeutic targets of interest in cancer and neurodegenerative disease. All KLKs appear to be secreted as inactive pro-forms (pro-KLKs) that are activated extracellularly by specific proteolytic release of their N-terminal pro-peptide. This processing is a key step in the regulation of KLK function. Much recent work has been devoted to elucidating the potential for activation cascades between members of the KLK family, with physiologically relevant KLK regulatory cascades now described in skin desquamation and semen liquefaction. Despite this expanding knowledge of KLK regulation, details regarding the potential for functional intersection of KLKs with other regulatory proteases are essentially unknown. To elucidate such interaction potential, we have characterized the ability of proteases associated with thrombostasis to hydrolyze the pro-peptide sequences of the KLK family using a previously described pro-KLK fusion protein system. A subset of positive hydrolysis results were subsequently quantified with proteolytic assays using intact recombinant pro-KLK proteins. Pro-KLK6 and 14 can be activated by both plasmin and uPA, with plasmin being the best activator of pro-KLK6 identified to date. Pro-KLK11 and 12 can be activated by a broad-spectrum of thrombostasis proteases, with thrombin exhibiting a high degree of selectivity for pro-KLK12. the results show that proteases of the thrombostasis family can efficiently activate specific pro-KLKs, demonstrating the potential for important regulatory interactions between these two major protease families.
- ItemSomente MetadadadosBaupain, A Plant Cysteine Proteinase That Hinders Thrombin-Induced Human Platelet Aggregation(Bentham Science Publ Ltd, 2012-04-01) Andrade, Sheila Siqueira [UNIFESP]; Silva, Mariana Cristina Cabral [UNIFESP]; Gouvea, Iuri Estrada [UNIFESP]; Kondo, Marcia Yuri [UNIFESP]; Juliano, Maria Aparecida [UNIFESP]; Sampaio, Misako Uemura [UNIFESP]; Oliva, Maria Luzia Vilela [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)Bauninia forficata is trivially known as cow paw, and popularly used in Brazil for treatment of diabetes mellitus. Denominated baupain a cysteine proteinase was purified from B. forficata leaves. In this study, we investigated the baupain effect on aggregation of isolated human platelets in vitro and the results show that baupain hinders thrombin - but not ADP-and collagen-induced platelet aggregation. With synthetic quenched-fluorescent peptides, the kinetics of the cleavage site of human proteinase-activated receptor 1 / 2 / 3 and 4 [PAR-1 / 2 / 3 and 4] by baupain was determined. In conclusion, similar to bromelain and papain, baupain hinders human platelets aggregation, probably through an unspecific cleavage in the Phe-Leu bond of PAR1.
- ItemAcesso aberto (Open Access)The hepatic clearance of recombinant tissue-type plasminogen activator decreases after an inflammatory stimulus(Associação Brasileira de Divulgação Científica, 2000-01-01) Nagaoka, Márcia Regina [UNIFESP]; Kouyoumdjian, Maria [UNIFESP]; Borges, Durval Rosa [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)We have shown that tissue-type plasminogen activator (tPA) and plasma kallikrein share a common pathway for liver clearance and that the hepatic clearance rate of plasma kallikrein increases during the acute-phase (AP) response. We now report the clearance of tPA from the circulation and by the isolated, exsanguinated and in situ perfused rat liver during the AP response (48-h ex-turpentine treatment). For the sake of comparison, the hepatic clearance of a tissue kallikrein and thrombin was also studied. We verified that, in vivo, the clearance of 125I-tPA from the circulation of turpentine-treated rats (2.2 ± 0.2 ml/min, N = 7) decreases significantly (P = 0.016) when compared to normal rats (3.2 ± 0.3 ml/min, N = 6). The AP response does not modify the tissue distribution of administered 125I-tPA and the liver accounts for most of the 125I-tPA (>80%) cleared from the circulation. The clearance rate of tPA by the isolated and perfused liver of turpentine-treated rats (15.5 ± 1.3 µg/min, N = 4) was slower (P = 0.003) than the clearance rate by the liver of normal rats (22.5 ± 0.7 µg/min, N = 10). After the inflammatory stimulus and additional Kupffer cell ablation (GdCl3 treatment), tPA was cleared by the perfused liver at 16.2 ± 2.4 µg/min (N = 5), suggesting that Kupffer cells have a minor influence on the hepatic tPA clearance during the AP response. In contrast, hepatic clearance rates of thrombin and pancreatic kallikrein were not altered during the AP response. These results contribute to explaining why the thrombolytic efficacy of tPA does not correlate with the dose administered.
- ItemSomente MetadadadosHigh-resolution structure of a Kazal-type serine protease inhibitor from the dengue vector Aedes aegypti(Int Union Crystallography, 2017) Torquato, Ricardo J. S. [UNIFESP]; Lu, Stephen [UNIFESP]; Martins, Nadia Helena; Tanaka, Aparecida S. [UNIFESP]; Barbosa Pereira, Pedro JoseBlood-feeding exoparasites are rich sources of protease inhibitors, and the mosquito Aedes aegypti, which is a vector of Dengue virus, Yellow fever virus, Chikungunya virus and Zika virus, is no exception. AaTI is a single-domain, noncanonical Kazal-type serine proteinase inhibitor from A. aegypti that recognizes both digestive trypsin-like serine proteinases and the central protease in blood clotting, thrombin, albeit with an affinity that is three orders of magnitude lower. Here, the 1.4 angstrom resolution crystal structure of AaTI is reported from extremely tightly packed crystals (similar to 22% solvent content), revealing the structural determinants for the observed inhibitory profile of this molecule.
- ItemSomente MetadadadosSimultaneous isolation of platelet factor 4 and glycoprotein IIb-IIIa complex from rabbit platelets, and characterization of specific chicken antibodies to assay them(Elsevier B.V., 2004-01-01) Santoro, M. L.; Barbaro, K. C.; Rocha, TRF da; Torquato, RJS; Hirata, I. Y.; Sano-Martins, I. S.; Inst Butantan; Pro Sangue Fdn; Universidade Federal de São Paulo (UNIFESP)Rabbits are frequently used as models for studying coagulation and platelet disorders. However, few reports on literature have dealt with the purification and characterization of rabbit platelet proteins. Herein a protocol for the simultaneous purification of rabbit platelet factor 4 (PF4) and platelet glycoprotein IIb-IIIa (GPIIb-IIIa, integrin alpha(IIb)beta(3)) is described. Specific antibodies were raised in laying chicken, which were used for assaying PF4 by ELISA, and GPIIb-IIIa by direct immunofluorescence and flow cytometry. Furthermore, the binding of monoclonal antibodies specific for GPIIb-IIIa complex (P2), ligand-induced binding site of GPIIIa (LIBS1) and rabbit P-selectin (12A7), as well as of polyclonal IgY specific for rabbit GPIIb-IIIa, was compared in quiescent and thrombin-activated platelets. Polyclonal anti-rabbit PF4 IgY was a specific and sensitive probe that could be used for assaying PF4 in plasma samples. GPIIb-IIIa expression was increased in thrombin-activated platelets, as evaluated by flow cytometric analysis using P2 and polyclonal antibodies raised in chickens. Rabbit GPIIb-IIIa also exhibited a conformational modification that caused the appearance of ligand-induced binding sites. Increased P-selectin expression, used as a positive control, was also noticeable in thrombin-activated platelets. These data evidence that antibodies raised in laying chickens specific to rabbit PF4 and GPIIb-IIIa, as well as certain monoclonal antibodies specific for human GPIIb-IIIa, may be used for investigating rabbit platelet physiology. (C) 2003 Elsevier B.V. All rights reserved.