Mercapturic acid formation in cultured opossum kidney cells

We investigated the last step of mercapturic acid formation, the N-acetylation of cysteine S-conjugates, in the established opossum kidney (OK) cell line which exhibits characteristics of the proximal tubule. S-Benzyl-L-cysteine was used as a model substance for such a cysteine S-conjugate. We succeeded in showing that OK cells absorb S-benzyl-L-cysteine via an active transport system which is inhibitable by phenylalanine. This transport follows Michaelis-Menten kinetics and the two characterizing parameters were determined: the Michaelis-Menten constant Km = 1.8 mmol/l, and the maximum of the difference between the intracellular and the extracellular concentration of S-benzyl-L-cysteine delta Cmax = 19.4 mmol/l. S-Benzyl-L-cysteine is converted to N-acetyl-S-benzyl-L-cysteine at a constant rate, which is independent of the extracellular S-benzyl-L-cysteine concentration. Under the tested experimental conditions this is probably due to saturation of the microsomal N-acetyltransferase catalyzing this reaction. In conclusion, we have shown that OK cells are a suitable model for studying mercapturate formation. They take up S-benzyl-L-cysteine mainly via the same carrier as phenylalanine, which is known to be transported in the rat by the high-capacity, low-affinity neutral amino acid carrier, and convert it to N-acetyl-L-benzyl-S-cysteine.