Isolation from rat kidney of a cytosolic high molecular weight cysteine-S-conjugate β-lyase with activity toward leukotriene E4

Abstract

A cytosolic high Mr cysteine-S-conjugate β-lyase (apparent Mr of ∼330,000) has been partially purified from rat kidneys. The high Mr lyase is also present in the mitochondria. The purified enzyme contains at least two proteins with apparent Mr values of ∼50,000 and ∼70,000. Activity is stimulated by dithiothreitol, α-keto acids, and pyridoxal 5′-phosphate; aminooxyacetate is an inhibitor. The enzyme catalyzes a competing (half) transamination reaction between pyridoxal 5′-phosphate cofactor and cysteine-S-conjugate substrate; added α-keto acids promote conversion of active site pyridoxamine 5′-phosphate to pyridoxal 5′-phosphate. The enzyme also catalyzes a full (but weak) transamination between L-phenylalanine and α-keto-γ-methiolbutyrate. The purified enzyme is not recognized by polyclonal rabbit antibodies to cytosolic rat kidney glutamine transaminase K (another cysteine-S-conjugate β-lyase of rat kidney) and has no obvious similarities to other pyridoxal 5′-phosphate-containing enzymes. In addition to catalyzing elimination reactions with S-(1,2-dichlorovinyl)-L-cysteine and S-(1,1,2,2-tetrafluoroethyl)-L-cysteine, the enzyme reacts with leukotriene E4 and 5′-S-cysteinyldopamine. Finally, the cytosolic and mitochondrial enzymes are activated by α-ketoglutarate. Thus, the possibility must be considered that, in kidneys of animals exposed to various cysteine conjugates, the high Mr lyase contributes to the generation of pyruvate, ammonia, and reactive fragments in vivo. Many cysteine conjugates are nephrotoxic, and the high Mr lyase(s) may be involved.