Methods for measuring cysteine S-conjugate β-lyase activity.

The cysteine conjugate β-lyase represents activities in cytoplasm and mitochondria catalyzed by at least eleven pyridoxal 5'-phosphate (PLP)-dependent enzymes in various tissues. These enzymes mediate bioactivation of cysteine S-conjugates of several haloalkanes and haloalkenes. The reaction occurs through either a direct β-elimination or a transamination followed by a retro-Michael rearrangement, resulting in the cleavage of a C-S bond. The resultant product is a reactive thiolate that rearranges to form thioacylating species. This unit presents several protocols for the assay of β-lyase activity and includes measurements of product formation and substrate loss as well as fluorescent activity stains. Support protocols describe the synthesis and high-performance liquid chromatography analysis of selected cysteine S-conjugates. Because of the diversity of enzymes that can catalyze a β-lyase reaction, each of the assays presented here may indicate only a portion of the potential β-lyase activity in a given biological preparation.