J Biol Chem, Vol. 274, Issue 4, 1994-1999, January 22, 1999

Leukotriene C₄ Is a Tight-binding Inhibitor of Microsomal Glutathione Transferase-1
EFFECTS OF LEUKOTRIENE PATHWAY MODIFIERS

Gerard Bannenberg, Sven-Erik Dahlén^¶, Marjanka Luijerink, Gerd Lundqvist, and Ralf Morgenstern

From the Institute of Environmental Medicine,  Division of Biochemical Toxicology and ^¶ Division of Experimental Asthma and Allergy Research, Karolinska Institutet, Box 210, SE-17177 Stockholm, Sweden

Microsomal glutathione transferase-1 (MGST-1) is an abundant protein that catalyzes the conjugation of electrophilic compounds with glutathione, as well as the reduction of lipid hydroperoxides. Here we report that leukotriene C₄ is a potent inhibitor of MGST-1. Leukotriene C₄ was found to be a tight-binding inhibitor, with a K_i of 5.4 nM for the unactivated enzyme, and 9.2 nM for the N-ethylmaleimide activated enzyme. This is the first tight-binding inhibitor characterized for this enzyme. Leukotriene C₄ was competitive with respect to glutathione and non-competitive toward the second substrate, CDNB. Analysis of stoichiometry supports binding of one molecule of inhibitor per homotrimer. Leukotrienes A₄, D₄, and E₄ were much weaker inhibitors of the purified enzyme (by at least 3 orders of magnitude). Leukotriene C₄ analogues, which have been developed as antagonists of leukotriene receptors, were found to display varying degrees of inhibition of MGST-1. In particular, the cysteinyl-leukotriene analogues SKF 104,353, ONO-1078, and BAYu9773 were strong inhibitors (IC₅₀ values: 0.13, 3.7, and 7.6 µM, respectively). In view of the partial structural similarity between MGST-1, leukotriene C₄ synthase, and 5-lipoxygenase activating protein (FLAP), it was of interest that leukotriene C₄ synthesis inhibitors (which antagonize FLAP) also displayed significant inhibition (e.g. IC₅₀ for BAYx1005 was 58 µM). In contrast, selective 5-lipoxygenase inhibitors such as zileuton only marginally inhibited activity at high concentrations (500 µM). Our discovery that leukotriene C₄ and drugs developed based on its structure are potent inhibitors of MGST-1 raises the possibility that MGST-1 influences the cellular processing of leukotrienes. These findings may also have implications for the effects and side-effects of drugs developed to manipulate leukotrienes.