Sulphonation of N-hydroxy-2-acetylaminofluorene by human dehydroepiandrosterone sulphotransferase.
Sulphonation of N-hydroxy-2-acetylaminofluorene by human dehydroepiandrosterone sulphotransferase.
复制标题
N-羟基-2-乙酰氨基芴被人脱氢表雄酮磺基转移酶磺化。
DOI：
10.1080/004982500237640
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发表时间：
2000
期刊：
影响因子：
--
通讯作者：
Walle,T
中科院分区：
文献类型：
--
作者：
Lewis,AJ;Otake,Y;Walle,UK;Walle,T
1. 目的是确定哪种人重组磺基转移酶 (ST) 同工型负责磺化，从而进一步潜在地激活经典的肝癌原癌物质 N-羟基-2-乙酰氨基芴 (N-OH-2AAF)。 2. N-OH-2AAF 与共底物 3'-磷酸腺苷-5'-磷酸硫酸酯 (PAPS) 以及人肝细胞溶胶或重组 P 型苯酚磺基转移酶 (P-PST)、M 型 PST、脱氢表雄酮-ST (DHEA-ST) 或雌激素 ST (EST) 一起孵育。通过 HPLC 测量从 PAPS 形成 3'-磷酸腺苷-5'-磷酸 (PAP) 的情况，用作测定磺基缀合率的测定方法。 3. 与基线水平相比，在 200 muM N-OH-2AAF 存在下，肝胞质中的 PAP 形成增加了 100% (p < 0.01)，对应于 19 pmol/min/mg 蛋白质的速率。然而，重组 P-PST 没有效果。这与之前使用粗酶制剂的建议形成对比。与 P-PST 一样，重组 M-PST 和 EST 不会磺化 N-OH-2AAF。另一方面，与基线值相比，在 200 μM N-OH-2AAF 存在下，重组 DHEA-ST 的 PAP 形成增加了 161% (p < 0.001)。 4. DHEA-ST 和人肝细胞质对 N-OH-2AAF 磺化的动力学研究给出了相似的表观 Kms。有趣的是，DHEA-ST 磺化 N-OH-2AAF 的 Vmax 与 DHEA（DHEA-ST 的天然底物）的 Vmax 非常相似。 5. 这是第一篇证明人类 DHEA-ST 参与 N-羟基化芳香酰胺致癌物磺化的论文。
1. The aim was to determine which human recombinant sulphotransferase (ST) isoform(s) were responsible for the sulphonation and, thus, potential further bioactivation of the classical hepatic procarcinogen N-hydroxy-2-acetylaminofluorene (N-OH-2AAF). 2. N-OH-2AAF was incubated together with the cosubstrate 3'-phosphoadenosine-5'- phosphosulphate (PAPS) and either human liver cytosol or recombinant P-form phenolsulphotransferase (P-PST), M-form PST, dehydroepiandrosterone-ST (DHEA-ST) or oestrogen ST (EST). Formation of 3'-phosphoadenosine-5'-phosphate (PAP) from PAPS, measured by HPLC, was used as the assay for determination of sulphoconjugation rates. 3. The liver cytosol produced a 100% increase in PAP formation in the presence of 200 muM N-OH-2AAF as compared with baseline levels (p < 0.01), corresponding to a rate of 19 pmol/min/mg protein. Recombinant P-PST, however, was without effect. This is in contrast to previous suggestions using crude enzyme preparations. Like P-PST, recombinant M-PST and EST did not sulphonate N-OH-2AAF. On the other hand, recombinant DHEA-ST produced a 161% increase in PAP formation in the presence of 200 muM N-OH-2AAF as compared with baseline values (p < 0.001). 4. Kinetic studies of N-OH-2AAF sulphonation by DHEA-ST and human liver cytosol gave similar apparent Kms. Interestingly, the Vmaxfor N-OH-2AAF sulphonation by DHEA-ST was very similar to that of DHEA, the natural substrate for DHEA-ST. 5. This is the first paper to demonstrate the involvement of the human DHEA-ST in the sulphonation of an N-hydroxylated aromatic amide carcinogen.