Protein arginine phosphorylation (pArg) is a relatively novel posttranslational modification. Protein arginine phosphatase YwlE negatively regulates arginine phosphorylation and consequently induces the expression of stress-response genes that are crucial for bacterial stress tolerance and pathogenic homolog Staphylococcus aureus virulence. However, little is known about the factors that affect the enzymatic activity of YwlE with the exception of the effect of oxidative stress. Herein, based on the hydrolysis of the chromogenic substrate p-nitrophenyl phosphate (pNPP) by YwlE, we investigate the role of metal cations and oxyanions in the regulation of YwlE activity. Interestingly, among the various cations that we tested, Ca²⁺ activates YwlE, while other cations, including Ag⁺, Co²⁺, Cd²⁺, and Zn²⁺, are inhibitory. Furthermore, as chemical analogues of phosphate, oxyanions play multiple roles in phosphatase activity. The regulatory switch Cys within the catalytic site regulates YwlE activity. Specifically, the thiol of this Cys could be alkylated by IAM (iodoacetamide) or oxidized by H₂O₂, resulting in enzymatic inhibition. Conversely, reducing reagents, such as DTT (dithiothreitol), β-me (β-mercaptoethanol), and TCEP (tris(2-carboxyethyl)phosphine) enhance YwlE activity. Additionally, as a stable analogue to pArg, pAIE binds to YwlE with a K_d of 149.1 nM and a binding stoichiometry n of 1.2 and inhibits YwlE with an IC₅₀ of 316.3 ± 12.73 μM. The inhibition and activation of YwlE may have broad implications for the physiology, pharmacology and toxicology of metal cations and oxyanions.

蛋白精氨酸磷酸化（pArg）是一种相对新颖的翻译后修饰。蛋白精氨酸磷酸酶YwlE负调节精氨酸磷酸化，因此诱导了应激反应基因的表达，这对于细菌的耐压力和致病性同源金黄色葡萄球菌毒力至关重要。然而，除了氧化应激的影响外，关于影响YwlE酶活性的因素知之甚少。本文中，基于YwlE水解发色底物对硝基苯基磷酸酯（p NPP），我们研究了金属阳离子和氧阴离子在YwlE活性调节中的作用。有趣的是，在我们测试的各种阳离子中，Ca ²⁺激活YwlE，而其他阳离子（包括Ag ⁺，Co 2 ⁺，Cd ²⁺和Zn ²⁺）具有抑制作用。此外，作为磷酸盐的化学类似物，氧阴离子在磷酸酶活性中起多种作用。催化位点内的调节开关Cys调节YwlE活性。具体地，该Cys的硫醇可以被IAM（碘乙酰胺）烷基化或被H ₂ O ₂氧化，从而导致酶促抑制。相反，还原剂，例如DTT（二硫苏糖醇），β-me（β-巯基乙醇）和TCEP（三（2-羧乙基）膦）可增强YwlE活性。另外，作为pArg的稳定类似物，pAIE的K _d与YwlE结合。149.1 nM的α和1.2的结合化学计量n抑制YwlE，IC ₅₀为316.3± 12.73μM 。YwlE的抑制和活化可能对金属阳离子和氧阴离子的生理学，药理学和毒理学具有广泛的意义。