Synthesis of 2-oxoglutarate derivatives and their evaluation as cosubstrates and inhibitors of human aspartate/asparagine-β-hydroxylase.
Synthesis of 2-oxoglutarate derivatives and their evaluation as cosubstrates and inhibitors of human aspartate/asparagine-β-hydroxylase.
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2-氧气谷物衍生物的合成及其作为人天冬氨酸/天冬酰胺-β-羟化酶的蛋白质和抑制剂的评估。
DOI：
10.1039/d0sc04301j
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发表时间：
2020-12-07
期刊：
影响因子：
8.4
通讯作者：
Schofield CJ
中科院分区：
文献类型：
--
作者：
Brewitz L;Nakashima Y;Schofield CJ
2-氧代戊二酸（2 OG）参与生物过程，包括由2 OG加氧酶催化的氧化，它是其共底物。真核生物2 OG加氧酶在胶原生物合成、脂质代谢、DNA/RNA修饰、转录调节和缺氧反应中发挥作用。天冬氨酸/天冬酰胺-β-羟化酶（AspH）是一种人2 OG加氧酶，催化内质网中表皮生长因子样结构域（EGFD）中Asp/Asn残基的翻译后羟化。AspH是化学兴趣，因为它的Fe（II）辅因子是由两个，而不是典型的三个残基复合。AspH在缺氧中上调，是癌细胞表面的预后标志物。我们描述了其天然2 OG辅助底物的衍生物如何调节AspH活性的研究。报道了一种有效的合成C3-和/或C4-取代的2 OG衍生物的方法，该方法通过氰基硫酰亚胺中间体进行。使用>30个2 OG衍生物进行的基于质谱的AspH测定显示，如晶体学和溶液分析所证明的，一些2 OG衍生物通过与2 OG竞争而有效地抑制AspH。其他2 OG衍生物可以取代2 OG，使底物羟基化。结果表明，细微的变化，如甲基到乙基取代，可以显着改变催化和抑制之间的平衡。3-甲基-2OG是一种存在于人类营养中的天然产物，是鉴定的最有效的替代共底物;晶体学分析揭示了（R）-3-甲基-2OG和其他2 OG衍生物与AspH的结合模式，并提供了周转和抑制之间的平衡。结果将使使用2 OG衍生物作为其他2 OG利用酶的机械探针，并建议2-含氧酸以外的2 OG可以采用一些2 OG加氧酶在体内。2-酮戊二酸衍生物的有效合成使得能够鉴定人天冬氨酸/天冬酰胺-β-羟化酶的抑制剂和新的共底物，如通过光谱学和晶体学研究所揭示的。
2-Oxoglutarate (2OG) is involved in biological processes including oxidations catalyzed by 2OG oxygenases for which it is a cosubstrate. Eukaryotic 2OG oxygenases have roles in collagen biosynthesis, lipid metabolism, DNA/RNA modification, transcriptional regulation, and the hypoxic response. Aspartate/asparagine-β-hydroxylase (AspH) is a human 2OG oxygenase catalyzing post-translational hydroxylation of Asp/Asn-residues in epidermal growth factor-like domains (EGFDs) in the endoplasmic reticulum. AspH is of chemical interest, because its Fe(ii) cofactor is complexed by two rather than the typical three residues. AspH is upregulated in hypoxia and is a prognostic marker on the surface of cancer cells. We describe studies on how derivatives of its natural 2OG cosubstrate modulate AspH activity. An efficient synthesis of C3- and/or C4-substituted 2OG derivatives, proceeding via cyanosulfur ylid intermediates, is reported. Mass spectrometry-based AspH assays with >30 2OG derivatives reveal that some efficiently inhibit AspH via competing with 2OG as evidenced by crystallographic and solution analyses. Other 2OG derivatives can substitute for 2OG enabling substrate hydroxylation. The results show that subtle changes, e.g. methyl- to ethyl-substitution, can significantly alter the balance between catalysis and inhibition. 3-Methyl-2OG, a natural product present in human nutrition, was the most efficient alternative cosubstrate identified; crystallographic analyses reveal the binding mode of (R)-3-methyl-2OG and other 2OG derivatives to AspH and inform on the balance between turnover and inhibition. The results will enable the use of 2OG derivatives as mechanistic probes for other 2OG utilizing enzymes and suggest 2-oxoacids other than 2OG may be employed by some 2OG oxygenases in vivo. Efficient synthesis of 2-oxoglutarate derivatives enables identification of inhibitors and novel cosubstrates of human aspartate/asparagine-β-hydroxylase as revealed by spectroscopic and crystallographic studies.
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影响因子：
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