气候变化领域集成服务门户(CCPortal): Short-lived intermediate in N2O generation by P450 NO reductase captured by time-resolved IR spectroscopy and XFEL crystallography

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DOI	10.1073/pnas.2101481118
	Short-lived intermediate in N2O generation by P450 NO reductase captured by time-resolved IR spectroscopy and XFEL crystallography
	Nomura T.; Kimura T.; Kanematsu Y.; Yamada D.; Yamashita K.; Hirata K.; Ueno G.; Murakami H.; Hisano T.; Yamagiwa R.; Takeda H.; Gopalasingam C.; Kousaka R.; Yanagisawa S.; Shoji O.; Kumasaka T.; Yamamoto M.; Takano Y.; Sugimoto H.; Tosha T.; Kubo M.; Shiro Y.
发表日期	2021
ISSN	0027-8424
卷号	118 期号:21
英文摘要	Nitric oxide (NO) reductase from the fungus Fusarium oxysporum is a P450-type enzyme (P450nor) that catalyzes the reduction of NO to nitrous oxide (N2O) in the global nitrogen cycle. In this enzymatic reaction, the heme-bound NO is activated by the direct hydride transfer from NADH to generate a short-lived intermediate (I), a key state to promote N–N bond formation and N–O bond cleavage. This study applied time-resolved (TR) techniques in conjunction with photolabile-caged NO to gain direct experimental results for the characterization of the coordination and electronic structures of I. TR freeze-trap crystallography using an X-ray free electron laser (XFEL) reveals highly bent Fe–NO coordination in I, with an elongated Fe–NO bond length (Fe–NO = 1.91 Å, Fe–N–O = 138°) in the absence of NAD+. TR-infrared (IR) spectroscopy detects the formation of I with an N–O stretching frequency of 1,290 cm−1 upon hydride transfer from NADH to the Fe3+–NO enzyme via the dissociation of NAD+ from a transient state, with an N–O stretching of 1,330 cm−1 and a lifetime of ca. 16 ms. Quantum mechanics/ molecular mechanics calculations, based on these crystallographic and IR spectroscopic results, demonstrate that the electronic structure of I is characterized by a singly protonated Fe3+–NHO•− radical. The current findings provide conclusive evidence for the N2O generation mechanism via a radical–radical coupling of the heme nitroxyl complex with the second NO molecule. © 2021 National Academy of Sciences. All rights reserved.
英文关键词	in crystallo microspectroscopy; Nitric oxide; NOR; time-resolved IR spectroscopy; XFEL-based crystallography
语种	英语
scopus关键词	ferric ion; heme; iron; nicotinamide adenine dinucleotide; nitric oxide reductase; nitrogen; nitrogen oxide; nitrous oxide; oxygen; P450 nitric oxide reductase; radical; reduced nicotinamide adenine dinucleotide; unclassified drug; Article; chemical bond; chemical structure; dissociation; enzyme mechanism; experimental study; in crystallo microspectroscopy; mathematical computing; molecular mechanics; quantum mechanics; spectroscopy; structure analysis; synthesis; time resolved near infrared spectroscopy; X ray crystallography; X ray free electron laser crystallography
来源期刊	Proceedings of the National Academy of Sciences of the United States of America
文献类型	期刊论文
条目标识符	http://gcip.llas.ac.cn/handle/2XKMVOVA/238909
作者单位	Graduate School of Life Science, University of Hyogo, Hyogo, 678-1297, Japan; RIKEN SPring-8 Center, Hyogo, 679-5148, Japan; Department of Chemistry, Graduate School of Science, Kobe University, Kobe, 657-8501, Japan; Graduate School of Information Sciences, Hiroshima City University, Hiroshima, 731-3194, Japan; Protein Crystal Analysis Division, Japan Synchrotron Radiation Research Institute, Hyogo, 679-5198, Japan; Department of Chemistry, Graduate School of Science, Nagoya University, Nagoya, 464-8602, Japan; Japan Science and Technology Agency, Core Research for Evolutional Science and Technology, Tokyo, 102-0075, Japan
推荐引用方式 GB/T 7714	Nomura T.,Kimura T.,Kanematsu Y.,et al. Short-lived intermediate in N2O generation by P450 NO reductase captured by time-resolved IR spectroscopy and XFEL crystallography[J],2021,118(21).
APA	Nomura T..,Kimura T..,Kanematsu Y..,Yamada D..,Yamashita K..,...&Shiro Y..(2021).Short-lived intermediate in N2O generation by P450 NO reductase captured by time-resolved IR spectroscopy and XFEL crystallography.Proceedings of the National Academy of Sciences of the United States of America,118(21).
MLA	Nomura T.,et al."Short-lived intermediate in N2O generation by P450 NO reductase captured by time-resolved IR spectroscopy and XFEL crystallography".Proceedings of the National Academy of Sciences of the United States of America 118.21(2021).