L-citrulline production from L-arginine by macrophage nitric oxide synthase. The ureido oxygen derives from dioxygen.
L-citrulline production from L-arginine by macrophage nitric oxide synthase. The ureido oxygen derives from dioxygen.
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DOI：
10.1016/s0021-9258(18)77366-3
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发表时间：
1990-08
期刊：
影响因子：
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通讯作者：
Nyoun Soo Kwon;Carl Nathan;C. Gilker;Owen W. Griffith;Dwight E. Matthews;D. Stuehr
中科院分区：
文献类型：
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作者：
Nyoun Soo Kwon;Carl Nathan;C. Gilker;Owen W. Griffith;Dwight E. Matthews;D. Stuehr
L精氨酸由L精氨酸经巨噬细胞一氧化氮(NO.)合成酶涉及精氨酸的水解或中间体的水合，因此预测了L-瓜氨酸中的水、氧的结合。巨噬细胞NO.合成酶与L精氨酸、NADPH、四氢生物蝶呤、FAD和二硫苏糖醇在H2(18)/16O2中孵育。用气相色谱/质谱法对反应生成的L瓜氨酸进行了分析。它的质谱图与在H_2(16)O/16O_2中产生的L-瓜氨酸的质谱图相匹配。以L-[Guanidino-13C]精氨酸为底物，证明m/z 99的碱基片段离子中含有缩脲基团。当酶在H_2(16)O/18O_2中反应时，以L-[胍-12C]精氨酸为底物的碱基片段离子移至m/z101，而以L-[胍-13C]精氨酸为底物的碱基片段离子移至m/z102。这些结果表明，巨噬细胞一氧化氮合酶L-瓜氨酸产物的缩脲基氧来源于氧气，而不是水。
Previously proposed mechanisms for the production of L-citrulline from L-arginine by macrophage nitric oxide (NO.) synthase involve either hydrolysis of arginine or hydration of an intermediate and thus predict incorporation of water oxygen into L-citrulline. Macrophage NO. synthase was incubated with L-arginine, NADPH, tetrahydrobiopterin, FAD, and dithiothreitol in H2(18)/16O2. L-Citrulline produced in this reaction was analyzed with gas chromatography/mass spectrometry. Its mass spectrum matched that of L-citrulline generated in H2(16)O/16O2. The base fragment ion of m/z 99 was shown to contain the ureido carbonyl group by using L-[guanidino-13C]arginine as substrate. When the enzyme reaction was performed in H2(16)O/18O2, the base fragment ion shifted to m/z 101 with L-[guanidino-12C]arginine as the substrate and to m/z 102 with L-[guanidino-13C]arginine. These results indicate that the ureido oxygen of the L-citrulline product of macrophage NO.synthase derives from dioxygen and not from water.