[Kinetics and mechanism of inactivation and reactivation of immobilized luciferase from fireflies Luciola mingrelica and the role of sulfhydryl in these processes].
The role of SH-groups of the enzyme in the catalytic and inactivation processes of soluble and immobilized on Sepharose 4B luciferase from the fireflies Luciola mingrelica was studied. A significant role of SH-groups of the soluble and immobilized enzyme was established by titrating the enzyme with 5,5'-dithiobis-2-nitrobenzoic acid. It was shown that the rate of inactivation of immobilized luciferase is limited by oxidation of the SH-groups of the enzyme; therefore dithiothreitol effectively protects the immobilized enzyme against inactivation. The kinetics of inactivation and reactivation of immobilized luciferase in the presence of dithiothreitol were studied. Dithiothreitol had no protective effect on soluble luciferase, which is indicative of another limiting step of inactivation of soluble luciferase. A mechanism of inactivation of Luciola mingrelica luciferase is proposed. The kinetic pattern of these processes is postulated and the rate constants of individual steps of inactivation of luciferase immobilized on Sepharose 4B were determined.
D000818:Animals; D001517:Coleoptera / Q000201:enzymology*; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology*; D004229:Dithiothreitol / Q000494:pharmacology; D004789:Enzyme Activation; D004800:Enzymes, Immobilized / Q000378:metabolism*; D007700:Kinetics; D008156:Luciferases / Q000378:metabolism*; D009579:Nitrobenzoates / Q000494:pharmacology*; D013438:Sulfhydryl Compounds
D004800:Enzymes, Immobilized; D009579:Nitrobenzoates; D013438:Sulfhydryl Compounds; D004228:Dithionitrobenzoic Acid; D008156:Luciferases; D004229:Dithiothreitol
Kinetika i mekhanizm inaktivatsii i reaktivatsii immobilizovannoĭ liutsiferazy svetliakov Luciola mingrelica i rol' sul'fgidril'nykh grupp v etikh protsessakh.