【作者】馬茜茜; 李淼; 王賀祥
【Author】MA Qianqian; LI Miao; WANG HexiangState Key Laboratory for Agrobiotechnology and Department of Microbiology; China Agricultural University; Beijing 100193; China
【作者機構】中國農業大學生物學院農業生物技術國家重點實驗室
【摘要】 采用DEAE-cellulose,CM-cellulose和Q-Sepharose離子交換層析和Superdex 75凝膠過濾層析,從云芝(Coriolus versicolor)發酵液中純化得到了一種漆酶,SDS-PAGE電泳檢測其為單一蛋白,分子量為67 kDa,N-端部分氨基酸序列為GIGPVADLTITNAAV。該酶可以催化氧化多種底物,其中對ABTS[2,2,-azinobis-(3-ethylbenzothiazoline-6-sulphonate)]的催化作用最強。最適反應條件為40℃,pH 5.0,其活性可以被Cu2+激活,被Fe3+、Hg2+、Fe2+抑制。該酶對HIV-1逆轉錄酶、乳腺癌MCF7細胞和肝癌HepG2細胞均具有抑制作用,其IC50分別為1.5μmol/L、2.3μmol/L和4.4μmol/L。
【Abstract】 A laccase from Coriolus versicolor was purified by ion exchange chromatography(DEAE-cellulose,CM-cellulose,Q-Sepharose) and gel filtration(Superdex 75) using fast protein liquid chromatography(FPLC).The enzyme is a monomeric protein with a molecular mass of 67 kDa based on SDS-PAGE and an N-terminal amino acid sequence of GIGPVADLTITNAAV.Purified laccase oxidized a range of phenolic and non-phenolic substrates,and exhibited highest activity on 2,2-azinobis-(3-ethylbenzothiazoline-6-sulphonate)(ABTS).With ABTS as the substrate,the enzyme displayed pH and temperature optima of 5.0 and 40 ℃,respectively and was significantly inhibited by Fe3+、Hg2+、and Fe2+ but stimulated by Cu2+.C.versicolor laccase inhibited HIV-1 reverse transcriptase activity and the proliferation of MCF7 and HepG2 cancer cells with IC50 values of 1.5 μmol/L,2.3 μmol/L and 4.4 μmol/L,respectively.
【關鍵詞】 云芝; 漆酶; 分離純化
【Key words】 Coriolus versicolor; laccase; enzyme purification
【基金】科技部“十一五”支撐項目“京郊地區種養業廢棄資源循環利用技術集成研究與示范”(編號:2007BAD89B07);; 中國農業大學生物學理科基地能力提高項目(科研訓練,J0730639)的部分研究內容~~
【文獻出處】 食用菌學報,Acta Edulis Fungi,編輯部郵箱,2010年02期
【DOI】CNKI:SUN:SYJB.0.2010-02-009
【分類號】S567.31