Purification of alpha-sarcin and an antifungal protein from Aspergillus giganteus by blue sepharose CL-6B affinity chromatography.
A simple method for preparation of alpha-sarcin and an antifungal protein (AFP) from the mold Aspergillus giganteus MDH 18894 has been developed. alpha-Sarcin and AFP were purified simultaneously by carboxymethylcellulose 52 cation-exchange chromatography and Blue Sepharose CL-6B affinity chromatography. By this method, 4.2 mg of alpha-sarcin and 6.8 mg of AFP were obtained from 2 liters of medium. Compared with other purification methods such as gel-filtration chromatography, this procedure was simple and specific. The purified alpha-sarcin and AFP were homogeneous characterized on SDS-polyacrylamide gel. The enzymatic activity of several ribosome-inactivating proteins such as alpha-sarcin, trichosanthin, and cinnamomin was significantly inhibited by the dye Cibacron blue F3GA. In 50 microliter of reaction mixture, 10 microM of the dye could inhibit 50% activity of cinnamomin (7 x 10(-9) M), whereas 50% inhibition of the enzymatic activity of trichosanthin (7 x 10(-9) M) and alpha-sarcin (1 x 10(-7) M) required 100 and 50 microM of the dye, respectively.
Hao
J J
JJ
State Key Laboratory of Molecular Biology, Laboratory of Ribosome Research, Laboratory of Plant Molecular Genetics, Shanghai Institute of Biochemistry, Academia Sinica, 320 Yue-Yang Road, Shanghai, 200031, China.
Xu
Y z
Yz
Geng
C D
CD
Liu
W Y
WY
Wang
E d
Ed
Gong
Z Z
ZZ
Ulbrich
N
N
D020418:Algal Proteins; D000935:Antifungal Agents / Q000737:chemistry*; D001230:Aspergillus / Q000737:chemistry*; D002455:Cell Division / Q000187:drug effects; D002846:Chromatography, Affinity / Q000379:methods; D002852:Chromatography, Ion Exchange / Q000379:methods; D004722:Endoribonucleases / Q000737:chemistry*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005656:Fungal Proteins / Q000302:isolation & purification; D011506:Proteins / Q000378:metabolism; D012260:Ribonucleases / Q000737:chemistry; D054790:Ribosome Inactivating Proteins, Type 2; D012270:Ribosomes / Q000378:metabolism; D012685:Sepharose / Q000031:analogs & derivatives / Q000378:metabolism; D014227:Triazines / Q000494:pharmacology; D015978:Trichosanthin / Q000378:metabolism; D020172:Verticillium / Q000187:drug effects
D020418:Algal Proteins; D000935:Antifungal Agents; D004791:Enzyme Inhibitors; D005656:Fungal Proteins; D011506:Proteins; D054790:Ribosome Inactivating Proteins, Type 2; D014227:Triazines; C017842:cibacron blue-sepharose; C061930:cinnamomin, Phytophthora cinnamomi; C014801:alpha-sarcin; C012842:Cibacron Blue F 3GA; D015978:Trichosanthin; D012685:Sepharose; D004722:Endoribonucleases; D012260:Ribonucleases