Purification and biochemical characterization of Eumiliin from Euphorbia milii var. hislopii latex

被引：38

作者：

Fonseca, K. C. ^{[1
]}

Morais, N. C. G. ^{[1
]}

Queiroz, M. R. ^{[1
]}

Silva, M. C. ^{[1
]}

Gomes, M. S. ^{[1
]}

Costa, J. O. ^{[2
,3
]}

Mamede, C. C. N. ^{[1
]}

Torres, F. S. ^{[4
]}

Penha-Silva, N. ^{[2
]}

Beletti, M. E. ^{[1
]}

Canabrava, H. A. N. ^{[1
]}

Oliveira, F. ^{[1
]}

机构：

[1] Univ Fed Uberlandia, Inst Ciencias Biomed, BR-38400 Uberlandia, MG, Brazil

[2] Univ Fed Uberlandia, Inst Genet & Bioquim, BR-38400 Uberlandia, MG, Brazil

[3] Inst Fed Educ Ciencias & Tecnol Triangulo Mineiro, Campus Ituiutaba, MG, Brazil

[4] Univ Fed Minas Gerais, Inst Ciencias Biol, Belo Horizonte, MG, Brazil

来源：

PHYTOCHEMISTRY | 2010年 / 71卷 / 07期

关键词：

Euphorbia milii; Crown-of-Thorns; Euphorbiaceae; Cysteine protease; Eumiliin; CUCUMISIN-LIKE PROTEASE; THROMBIN-LIKE ENZYME; SERINE-PROTEASE; ENDEMIC AREA; ANTIPLATELET; PROTEINASE; COAGULANT; SARCOCARP; CLONING; VENOMS;

D O I：

10.1016/j.phytochem.2010.02.009

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

A protease, which we designate Eumiliin, was isolated from the latex of Euphorbia milii var. hislopii by a combination of ion-exchange chromatographic steps using DEAE-Sephacel and gel-filtration with Sephadex G-75. Eumiliin is a monomeric protein with an apparent molecular mass of 30 kDa by SDS-PAGE under reducing conditions and gave one main peak at 29,814 KDa in MALDI-TOF/TOF mass spectrometry. Eumiliin has caseinolytic and fibrinogenolytic activities, but no hemorrhagic or defibrinating activities. The enzyme readily hydrolyzes the A alpha-chain of fibrinogen and, more slowly, the B beta-chain. Its fibrinogenolytic activity is inhibited by beta-mercaptoethanol and leupeptin. In contrast, EDTA and benzamidine did not affect the activity of Eumiliin. The caseinolytic activity of Eumiliin had a pH optimum of 8.0 and was stable in solution at up to 40 degrees C; activity was completely lost at >= 80 degrees C. Intraplantar injection of Eumiliin (1-25 mu g/paw) caused a dose- and time-dependent hyperalgesia, which peaked 1-5 h after enzyme injection. Intraplantar injection of Eumiliin (1-25 mu g/paw) also caused an oedematogenic response that was maximal after 1 h. Morphological analyses indicated that Eumiliin induced an intense myonecrosis, with visible leukocyte infiltrate and damaged muscle cells 24 h after injection. (C) 2010 Elsevier Ltd. All rights reserved.

引用

页码：708 / 715

页数：8

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