Identification of a high affinity leukotriene C₄-binding protein in rat liver cytosol as glutathione S-transferase

A soluble high affinity binding unit for leukotriene (LT) C₄ in the high speed supernatant of rat liver homogenate was characterized at 4°C as having a single type of saturable affinity site with a dissociation constant of 0.77 ± 0.27 nM (mean ± S.E., n = 5). The binding activity was identified as the liver cytosolic subunit 1 (Ya) of glutathione S-transferase, commonly known as ligandin, by co-purification with the catalytic activity during DEAE-cellulose column chromatography and 11,12,14,15-tetrahydro-LTC₄ (LTC₂)-affinity gel column chromatography; resolution into two major bands by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of M(r) 23,000 and 25,000, of which only the smaller protein was labeled with [³H]LTC₄ coupled via a photoaffinity cross-linking reagent; and immunodiffusion analysis with rabbit antiserum to glutathione S-transferase which showed a line of identity between the purified LTC₄-binding protein and rat liver glutathione S-transferase. The affinity-purified binding protein bound 800 pmol of [³H]LTC₄/mg of protein and possessed 12 μmol/min/mg of glutathione transferase activity as assayed with 1-chloro-2,4-dinitrobenzene as substrate. The enzyme activity of the cytosolic LTC₄-binding protein was inhibited by submicromolar quantities of unlabeled LTC₄, and the binding activity for [³H]LTC₄ was blocked by the ligandin substrates, hematin and bilirubin. The high affinity interaction between LTC₄ and glutathione S-transferase suggests that glutathione S-transferase may have a role in LTC₄ disposition and that previous studies of LTC₄ binding to putative receptors in nonresponsive tissues may require redefinition of the binding unit.