Persistent Activation of cGMP-Dependent Protein Kinase by a Nitrated Cyclic Nucleotide via Site Specific Protein S-Guanylation
Persistent Activation of cGMP-Dependent Protein Kinase by a Nitrated Cyclic Nucleotide via Site Specific Protein S-Guanylation
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DOI：
10.1021/acs.biochem.5b00774
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发表时间：
2016-02-09
期刊：
影响因子：
2.9
通讯作者：
Akaike, Takaaki
中科院分区：
文献类型：
--
作者：
Akashi, Soichiro;Ahmed, Khandaker Ahtesham;Akaike, Takaaki
8-硝基鸟苷3',5'-环单磷酸（8-硝基cGMP）是鸟苷3',5'-环单磷酸（cGMP）的硝化衍生物，是在活性氧和一氧化氮的生产相关条件下内源性形成的。它在细胞信号传导调控中作为亲电第二信使，通过cGMP部分共价内聚到蛋白质硫醇（蛋白质s -胍基化）诱导氧化还原敏感蛋白硫醇的翻译后修饰。在这里，我们证明了8-硝基cGMP可能会s -鸟苷化cGMP依赖性蛋白激酶（PKG）的巯基，PKG是细胞内cGMP的主要受体蛋白之一，并控制各种细胞反应。发现PKG的s -胍基化以特定位点的方式发生；在11个Cys残基中，Cys42和Cys195是敏感残基。重要的是，Cys195的s -胍基化位于PKG的高亲和力cGMP结合区域，通过体外激酶测定和器官浴测定可以引起持续的酶激活。在体内，PKG的s -胍基化被证明在没有任何特异性处理的情况下发生在小鼠体内，并且脂多糖管理显著增强。这些发现为进一步研究s -胍基化依赖性持续PKG激活的生理和病理生理作用提供了依据。
8-Nitroguanosine 3',5'-cyclic monophosphate (8-nitro-cGMP) is a nitrated derivative of guanosine 3',5'-cyclic monophosphate (cGMP) formed endogenously under conditions associated with production of both reactive oxygen species and nitric oxide. It acts as an electrophilic second messenger in the regulation of cellular signaling by inducing a post-translational modification of redox-sensitive protein thiols via covalent adduction of cGMP moieties to protein thiols (protein S-guanylation). Here, we demonstrate that 8-nitro-cGMP potentially S-guanylates thiol groups of cGMP-dependent protein kinase (PKG), the enzyme that serves as one of the major receptor proteins for intracellular cGMP and controls a variety of cellular responses. S-Guanylation of PKG was found to occur in a site specific manner; Cys42 and Cys195 were the susceptible residues among 11 Cys residues. Importantly, S-guanylation at Cys195, which is located in the high-affinity cGMP binding domain of PKG, causes persistent enzyme activation as determined by in vitro kinase assay as well as by an organ bath assay. In vivo, S-guanylation of PKG was demonstrated to occur in mice without any specific treatment and was significantly enhanced by lipopolysaccharide administration. These findings warrant further investigation in terms of the physiological and pathophysiological roles of S-guanylation-dependent persistent PKG activation.