Rapid purification of 2,4-dichlorophenol hydroxylase by biospecific desorption from 10-carboxydecylamino-sepharose.
10-Carboxydecylamino-Sepharose, which bears a mixture of ionic and aliphatic substituent groups, adsorbs 2,4-dichlorophenol hydroxylase from Acinetobacter in a non-biospecific manner. The enzyme has been specifically desorbed by its substrate, 2,4-dichlorophenol, giving a 42-fold purification (to greater than 90% purity) in a single step. The enzyme contained 3.1 moles of FAD per mole and displayed a catalytic constant of 14.7 s(-1). Mixed-function adsorbents probably have wide applicability for biospecific desorption of proteins. The present report indicates that they may be useful in the purification of aromatic hydroxylases bearing flavin prosthetic groups that readily dissociate in conventional purification procedures employing conditions of high ionic strength.
D000150:Acinetobacter / Q000201:enzymology; D002846:Chromatography, Affinity / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000302:isolation & purification*; D012685:Sepharose / Q000031:analogs & derivatives
D005182:Flavin-Adenine Dinucleotide; C037346:10-carboxydecylamino-sepharose; D012685:Sepharose; D006899:Mixed Function Oxygenases; C034354:2,4-dichlorophenol hydroxylase