[Kinetic mechanisms of linoleic acid oxidation by 5-lipoxygenase from Solanum tuberosum L].
Linoleic acid oxidation by 5-lipoxygenase from Solanum tuberosum has been studied as affected by sodium dodecylsulfate (Ds-Na). The reaction system consisted of 5-lipoxygenase and mixed micelles of linoleic acid and Lubrol PX. It contained varying amounts of the enzyme effector--Ds-Na. The enzyme showed a pronounced cooperativity, and the reaction was governed by the Hill equation with h = 3.7. On the other side, increasing amounts of Ds-Na added to the system caused a tremendous increase of enzyme activity and simultaneous decline of h, with was proportional to Ds-Na concentration. Ds-Na had dual effect on 5-lipoxygenase--there was an optimal concentration of the compound (0.34 mM Lubrol PX; 0.2 mM LA; 0.13 mM Ds-Na; pH = 6.3) causing the 4-fold highest activation and h = 1.6. The further increase of Ds-Na led to the enzyme inhibition. If Ds-Na was 0.5 mM, h became 1. At this point, each molecule of 5-lipoxygenase bound 3 molecules of Ds-Na and 1 molecule of linoleic acid, thus the total number of occupied binding sites was 4. A kinetic scheme of 5-lipoxygenase reaction has been proposed. It was found that the enzyme's kinetic behaviour could be explaine if assumed an existence of a special noncatalytic binding centre capable of binding several (up to 3) molecules of either substrate, or effector. Such a centre can serve as an anchoring site facilitating the enzyme binding to the surface of lipid aggregates containing insolubilized substrate molecules. Replacing linoleic acid in the binding site, Ds-Na activates the enzyme, possibly due to the much more effective translocation of 5-lipoxygenase to the surface of lipid aggregates. This mechanism can be an universal alternative to the FLAP-type regulation of 5-lipoxygenase activities.
Kharchenko
O V
OV
Institute of Bioorganic Chemistry and Petrochemistry of National Academy of Sciences of Ukraine, Kyiv.
Kulinichenko
H I
HI
Butovych
I A
IA
D001094:Arachidonate 5-Lipoxygenase / Q000378:metabolism*; D001692:Biological Transport; D007700:Kinetics; D019787:Linoleic Acid / Q000378:metabolism*; D010084:Oxidation-Reduction; D012967:Sodium Dodecyl Sulfate; D011198:Solanum tuberosum / Q000201:enzymology*; D013379:Substrate Specificity
D012967:Sodium Dodecyl Sulfate; D019787:Linoleic Acid; D001094:Arachidonate 5-Lipoxygenase