High performance affinity chromatography of Bacillus neutral proteases
Eijsink, V.G.; van den Burg, B.; Venema, G.
Biotechnology and Applied Biochemistry 14(3): 275-283
1991
ISSN/ISBN: 0885-4513 PMID: 1777113 Document Number: 383063
Bacillus neutral proteases were purified using bacitracin-silica as an affinity medium. Several chromatographic procedures were investigated, including high speed runs on columns with 40- to 60-microns silica particles. The high speed procedure enabled the purification of 4.9 mg of B. subtilis neutral protease directly from 165-ml culture supernatant within 1.5 h. The neutral proteases of B. polymyxa and B. stearothermophilus were also purified. The latter enzyme was further concentrated by a second affinity chromatography step, using Sepharose with glycyl-D-phenylalanine as a ligand. During the purification procedures isopropanol was used to prevent autodigestion of the enzymes.