title: Purification of L-kynurenine 3-hydroxylase by affinity chromatography.
abstract: NADP immobilized on agarose is able to adsorb L-kynurenine 3-hydroxylase. The enzyme is released from the adsorbent by passage of a buffer containing 0.5 mM NADP through the column. L-Kynurenine 3-hydroxylase was purified 26-fold with a yield of 12% from mitochondrial outer membrane with a procedure involving DEAE-Sepharose CL-6B, Sephacryl S-200 chromatography and NADP-agarose affinity chromatography. This monooxygenase was a homogeneous protein, giving a monomeric molecular weight of 145,000, which had neither any significant NADPH diaphorase activity nor cytochrome b5-like haem protein. However, the enzyme did not show affinity for a column with L-kynurenine coupled to the gel with a suitable spacer group, and AMP did not serve as an effective ligand in an affinity resin.
mesh_terms: D000818:Animals; D002846:Chromatography, Affinity / Q000379:methods; D007425:Intracellular Membranes / Q000201:enzymology; D007737:Kynurenine / Q000302:isolation & purification; D050601:Kynurenine 3-Monooxygenase; D008930:Mitochondria, Liver / Q000201:enzymology*; D006899:Mixed Function Oxygenases / Q000302:isolation & purification*; D008970:Molecular Weight; D051381:Rats
chemical_list: D007737:Kynurenine; D006899:Mixed Function Oxygenases; D050601:Kynurenine 3-Monooxygenase