Biomolecule sulfation catalyzed by sulfotransferases (STases) plays a profound role in numerous biological processes. However, the exact biofunctions of the sulfation modifications still remain largely unknown partially because STases are difficult to assay. Most of the existing STase assays are targeted to discriminate the property changes of the sulfated substrate against the un‐sulfated ones, which typically suffer from several drawbacks due to the labile sulfated products, inhibitory effect of the 3'‐phosphoadenosine‐5'‐phosphate (PAP) by‐product as well as the lack of generality. To address such issues, herein, we have developed a versatile and general fluorescence turn‐on strategy for assaying STase activities. Unlike traditional STase assays, this work is targeted to measuring the phosphate ion (Pi) released from the enzymatic degradation of PAP by‐product during the sulfation reaction, which can lead to fluorescence enhancement of a calcein/Ce³⁺ system. Since PAP is the common by‐product of all kinds of STase reactions, this assay is universally applicable to the varying kinds of STases. In this regard, the intrinsic instability of the sulfated substrates will no longer influence the assay accuracy. Furthermore, the inhibitory effect of PAP on the STase reaction, which is common in traditional STase assays, are effectively removed in this study, thereby allowing more accurate determination of the enzyme activity.

中文翻译：

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通过钙黄绿素/ Ce3 +系统检测游离磷酸盐离子的通用荧光开启磺基转移酶测定

磺基转移酶（STases）催化的生物分子硫酸化在许多生物学过程中都起着重要作用。但是，硫酸盐修饰的确切生物功能仍然很大程度上未知，部分原因是STases难以测定。大多数现有的STase分析法的目标是区分硫酸化底物与未硫酸化底物的特性变化，这通常由于不稳定的硫酸化产物，3'-磷酸腺苷-5'-磷酸的抑制作用而遭受一些弊端。 （PAP）副产品以及缺乏通用性。为了解决这些问题，在本文中，我们开发了一种通用的通用荧光开启策略来检测STase活性。与传统的STase分析不同，³⁺系统。由于PAP是各种STase反应的常见副产物，因此该测定法普遍适用于各种STase。在这方面，硫酸化底物的固有不稳定性将不再影响测定的准确性。此外，这项研究有效地消除了PAP对STase反应的抑制作用，这在传统的STase分析中很常见，因此可以更准确地确定酶活性。