Shoeman D W, Shirota F N, DeMaster E G, Nagasawa H T
Medical Research Laboratories, VA Medical Center, Minneapolis, MN 55417 USA.
Alcohol. 2000 Jan;20(1):55-9. doi: 10.1016/s0741-8329(99)00056-7.
Nitroxyl (HNO) is the aldehyde dehydrogenase (AIDH) inhibitor produced by catalase action on cyanamide. Incubation of N-acetyl-L-cysteine (NAC), a reagent with a free sulfhydryl group, with Piloty's acid (a nitroxyl generator) suggested that NAC was acting as a competitive "trap" for nitroxyl. Elucidation of the structure of this reaction product should give an insight as to how nitroxyl interacts with AIDH, a sulfhydryl enzyme. We now present evidence that the product formed is N-acetyl-L-cysteinesulfinamide (NACS). We have synthesized NACS and showed that this synthetic product was identical to the product formed in the trapping experiment. Both had identical RT values by reverse phase HPLC and identical RF values by TLC using three different solvent systems. The structural identification of this nitroxyl trapped product as a sulfinamide now allows the chemical confirmation of the active-site cysteine residue of AIDH as Cys-302.
硝酰基（HNO）是过氧化氢酶作用于氰胺产生的醛脱氢酶（AIDH）抑制剂。具有游离巯基的试剂N - 乙酰 - L - 半胱氨酸（NAC）与皮洛蒂酸（一种硝酰基生成剂）一起孵育表明，NAC作为硝酰基的竞争性“捕获剂”起作用。阐明该反应产物的结构应能深入了解硝酰基如何与作为巯基酶的AIDH相互作用。我们现在提供证据表明形成的产物是N - 乙酰 - L - 半胱氨酸亚磺酰胺（NACS）。我们已经合成了NACS，并表明该合成产物与捕获实验中形成的产物相同。通过反相高效液相色谱法，二者具有相同的保留时间（RT）值；使用三种不同溶剂系统通过薄层层析法，二者具有相同的比移值（RF）。这种被硝酰基捕获的产物作为亚磺酰胺的结构鉴定现在允许化学确认AIDH的活性位点半胱氨酸残基为Cys - 302。
