2,6-Dichloro-4-nitrophenol (DCNP)-35sulfate was identified and quantified by an HPLC-radiometric assay following its biosynthesis in vitro from 35S-labeled 3'-phosphoadenosine-5'-phosphosulfate (PAP35S) by phenolsulfotransferase (PST) of rat liver cytosol. Acid hydrolysis of DCNP-35sulfate produced almost stoichiometric release of inorganic 35sulfate and DCNP. In two-substrate experiments of sulfation of p-nitrophenol (p-NP) or dopamine (prototype substrates for P and M human PST forms), 10 microM DCNP inhibited the reactions by about 15 and 78%, respectively. This contrasts with its action on PST of human origin where the P-PST was more sensitive to DCNP inhibition. In all mixed bi-substrate experiments, a reciprocal relationship between the two sulfated products was observed. Kinetic data showed that p-NP inhibited the sulfation of DCNP competitively. Likewise the sulfation of p-NP and dopamine was competitively inhibited by DCNP. However, non-competitive inhibition was observed in the sulfation of p-NP by DCNP, measured at varying concentrations of PAP35S. The above kinetic data suggest that DCNP is an alternate-substrate inhibitor of rat liver PST.

中文翻译：

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2,6-二氯-4-硝基苯酚（DCNP），苯酚磺基转移酶的另一种底物抑制剂

在通过35S标记的3'-磷酸腺苷-5'-磷酸硫酸酯（PAP35S）通过酚磺基转移酶（PST）体外生物合成后，通过HPLC放射分析法鉴定并定量了2,6-二氯-4-硝基苯酚（DCNP）-35硫酸酯大鼠肝细胞溶质。DCNP-35硫酸盐的酸水解产生了几乎化学计量的无机35硫酸盐和DCNP释放。在对硝基苯酚（p-NP）或多巴胺（P和M型人PST形式的原型底物）的硫酸盐化的两个底物实验中，10 microM DCNP分别抑制了反应约15％和78％。这与它对人源PST的作用相反，在PST中，P-PST对DCNP抑制更为敏感。在所有混合的双底物实验中，观察到两种硫酸化产物之间的相互关系。动力学数据表明，p-NP竞争性地抑制了DCNP的硫酸化。同样地，DCNP竞争性地抑制了对NP和多巴胺的硫酸化。但是，在不同浓度的PAP35S上测得的DCNP对p-NP的硫酸化作用中观察到非竞争性抑制作用。以上动力学数据表明DCNP是大鼠肝脏PST的另一种底物抑制剂。