Effect of nonionic detergents on lipoxygenase catalysis.
In many studies on lipoxygenase catalysis, nonionic detergents are used to obtain an optically transparent solution of the fatty acid substrate. In order to resolve some controversies that exist with regard to the interpretation of kinetic data obtained with solutions containing nonionic detergents, a systematic investigation was undertaken into the effects of Lubrol, Tween-20 and Triton X-100 (0-0.8 g/L) on the kinetics of linoleate (2.5-110 microM) dioxygenation, catalyzed by lipoxygenase-1 or lipoxygenase-2 from soybean, at pH 9 or 10, at 25 degrees C. Under most conditions, it was found that the detergents slowed down the reaction. However, at high linoleate concentrations, where substrate inhibition of lipoxygenase is significant, small amounts of detergent increased the dioxygenation rate. In a quantitative analysis of the results, a kinetic model in which the incorporation of linoleate in the detergent micelles is formulated as a simple reversible equilibrium, and in which both lipoxygenase-1 and -2 interact with free linoleate, but not with linoleate incorporated in the micelles, appeared to be sufficient to predict experimental results over a wide range of experimental conditions. According to this model, the changes in the dioxygenation kinetics caused by the presence of nonionic detergents are similar (but not equal) to those caused by competitive inhibitors. The conclusions that monomeric, nonmicellar linoleate is the preferred substrate for lipoxygenase and that the observed inhibition and stimulation are solely due to changes in the effective linoleate concentration strongly corroborate the earlier observations by Galpin and Allen [Biochim. Biophys. Acta 488 (1977), 392-401].
Schilstra
M J
MJ
Bijvoet Center for Biomolecular Research, Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Veldink
G A
GA
Vliegenthart
J F
JF
D002384:Catalysis; D003902:Detergents / Q000494:pharmacology*; D006863:Hydrogen-Ion Concentration; D007477:Ions; D007700:Kinetics; D019787:Linoleic Acid; D008041:Linoleic Acids / Q000378:metabolism; D008084:Lipoxygenase / Q000378:metabolism*; D017830:Octoxynol / Q000494:pharmacology; D011092:Polyethylene Glycols / Q000494:pharmacology; D011136:Polysorbates / Q000494:pharmacology; D013025:Glycine max / Q000201:enzymology
D003902:Detergents; D007477:Ions; D008041:Linoleic Acids; D011136:Polysorbates; D011092:Polyethylene Glycols; C005092:lubrol; D017830:Octoxynol; D019787:Linoleic Acid; D008084:Lipoxygenase