title: [Properties of beta-carotene-15,15'-dioxygenase, stabilized during purification with lutein and dithiothreitol].
abstract: Preparations of beta-carotene 15,15'-dioxygenase (EC 1.13.11.21) obtained with or without lutein and dithiothreitol (DTT) as stabilizers display similar properties. The stabilized preparation has an apparent Km of 17 microM and Vmax of 2.8 nM/h/mg of protein, a pH optimum of 8.1, is sensitive to SH-agents and is activated by Fe2+, DTT or glutathione used at optimal concentrations of 0.5 mM, 2 mM, and 5 mM respectively. At DTT concentration above 2 mM the enzyme activity decreases drastically, which testifies to the importance of maintaining the thiol and disulphide group ratio in the protein molecule at an optimal level. At 4 mM Fe2+ there occurs a nonenzymatic formation of apo-carotenals; however, retinal is formed only after addition of commensurate amounts of thiol compounds. The enzyme activity depends on the ionic strength of the incubation medium, the maximal effect being observed with 0.4 M K-phosphate buffer. Substitution of phosphate for Cl- partly inhibits the enzyme. The optimal temperature for this reaction is 45 degrees. A simultaneous use of optimal incubation parameters (2 mM DTT, 0.4 M K-phosphate buffer pH 8.1 and 45 degrees C) increases the retinal yield 2.5-fold against control.
authors: Iu V
authors: IuV
authors: Ershov
authors: A A
authors: AA
authors: Dmitrovskiĭ
authors: V Ia
authors: VIa
authors: Bykhovskiĭ
mesh_terms: D004229:Dithiothreitol / Q000737:chemistry*; D004789:Enzyme Activation; D004795:Enzyme Stability; D014975:Lutein / Q000737:chemistry*; D009994:Osmolar Concentration; D010105:Oxygenases / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D013696:Temperature; D050559:beta-Carotene 15,15'-Monooxygenase
chemical_list: D010105:Oxygenases; D050559:beta-Carotene 15,15'-Monooxygenase; D004229:Dithiothreitol; D014975:Lutein
vernacular_title: Svoĭstva beta-karotin-15,15'-dioksigenazy, stabilirzirovannoĭ v protsesse vydeleniia liutenom i ditiotreitolom.