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N-Glycosylation at Asn695 might suppress inducible nitric oxide synthase activity by disturbing electron transfer
Acta Biochimica et Biophysica Sinica ( IF 3.7 ) Pub Date : 2020-11-21 , DOI: 10.1093/abbs/gmaa132
Jianghong Yan 1, 2 , Fei-Fei Shang 1 , An He 3 , Shupeng Hu 3 , Suxin Luo 3 , Yong Xia 1, 3, 4
Affiliation  

Abstract
Inducible nitric oxide synthase (iNOS) plays critical roles in the inflammatory response and host defense. Previous research on iNOS regulation mainly focused on its gene expression level, and much less is known about the regulation of iNOS function by N-glycosylation. In this study, we report for the first time that iNOS is N-glycosylated in vitro and in vivo. Mass spectrometry studies identified Asn695 as an N-glycosylation site of murine iNOS. Mutating Asn695 to Gln695 yields an iNOS that exhibits greater enzyme activity. The essence of nitric oxide synthase catalytic reaction is electron transfer process, which involves a series of conformational changes, and the linker between the flavin mononucleotide-binding domain and the flavin adenine dinucleotide-binding domain plays vital roles in the conformational changes. Asn695 is part of the linker, so we speculated that attachment of N-glycan to the Asn695 residue might inhibit activity by disturbing electron transfer. Indeed, our NADPH consumption results demonstrated that N-glycosylated iNOS consumes NADPH more slowly. Taken together, our results indicate that iNOS is N-glycosylated at its Asn695 residue and N-glycosylation of Asn695 might suppress iNOS activity by disturbing electron transfer.


中文翻译:

Asn695处的N-糖基化可能通过干扰电子转移来抑制诱导型一氧化氮合酶活性

摘要
诱导型一氧化氮合酶(iNOS)在炎症反应和宿主防御中起关键作用。以前关于iNOS调控的研究主要集中在其基因表达水平上,而关于通过N-糖基化调控iNOS功能的了解还很少。在这项研究中,我们首次报道了iNOS在体外体内均被N-糖基化。质谱研究确定Asn 695为鼠iNOS的N-糖基化位点。将Asn 695突变为Gln 695产生具有更高酶活性的iNOS。一氧化氮合酶催化反应的实质是电子转移过程,它涉及一系列构象变化,黄素单核苷酸结合域和黄素腺嘌呤二核苷酸结合域之间的连接基在构象改变中起着至关重要的作用。Asn 695是连接子的一部分,因此我们推测N-聚糖与Asn 695残基的附着可能会通过干扰电子转移来抑制活性。确实,我们的NADPH消耗结果表明N-糖基化iNOS消耗NADPH的速度更慢。总之,我们的结果表明,iNOS的是在它的天冬酰胺N-糖基化695天冬酰胺的残留和N-糖基化695 可能通过干扰电子传递来抑制iNOS活性。
更新日期:2020-12-29
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