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The application of hollow fiber cartridges in the purification of antitumor antibiotic, neocarzinostatin from Streptomuces carzinostaticus
Author(s) -
Li HuangHsien,
Hariharan Parameswaran,
Chin DerHang
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.5.a630-b
Subject(s) - enediyne , neocarzinostatin , antibiotics , chemistry , chromatography , streptomyces , cartridge , ion exchange , bacteria , biochemistry , biology , stereochemistry , organic chemistry , materials science , dna , ion , metallurgy , genetics
Neocarzinostatin is a potent enediyne antibiotic chromoprotein, secreted by Streptomyces carzinostaticus. It consists of an 11 kDa all‐beta protein that is non‐covalently associated with a very labile 659 Da enediyne antibiotic chromophore. Although the production and purification of neocarzinostatin were reported in detail earlier, execution of the reported methods in our laboratory frequently results in very poor antibiotic yield because of its instability. Crude culture filtrates containing active neocarzinostatin antibiotic from fermentation of S. carzinostaticus (ATCC15945 and ATCC15944) were prepared using the reported procedures. Here we describe a slight modification to replace the clay‐involved precipitation step in the reported method of purifying the chromoprotein antibiotic neocarzinostatin. The modified procedure employs a quick concentrating step by involving hollow fiber cartridges to avoid extensive degradation of the labile antibiotic during the prolonged purification steps. A combination of 500 kDa (or 50 kDa) and 3 kDa cutoff cartridges is used to exclude large insoluble materials and to retain fractions containing active neocarzinostatin. Followed by quick concentrating step, buffer exchange with samples is then carried out using the 3 kDa cutoff cartridge. This allows the semi‐purified samples to be applied directly to ion‐exchange chromatographic purification. This work is funded by National Science Council and National Health Research Institute of Taiwan, R.O.C.