Open and closed structures of L-arginine oxidase by cryo-electron microscopy and X-ray crystallography.

IF 2.1 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY
Hiroki Yamaguchi, Kazutoshi Takahashi, Nobutaka Numoto, Hiroshi Suzuki, Moemi Tatsumi, Akiko Kamegawa, Kouki Nishikawa, Yasuhisa Asano, Toshimi Mizukoshi, Hiroshi Miyano, Yoshinori Fujiyoshi, Masayuki Sugiki
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引用次数: 0

Abstract

L-arginine oxidase (AROD, EC 1.4.3.25) is an oxidoreductase that catalyzes the deamination of L-arginine, with flavin adenine dinucleotide (FAD) as a cofactor. Recently identified AROD from Pseudomonas sp. TPU 7192 (PT-AROD) demonstrates high selectivity for L-arginine. This enzyme is useful for accurate assays of L-arginine in biological samples. The structural characteristics of the FAD-dependent AROD, however, remain unknown. Here, we report the structure of PT-AROD at a resolution of 2.3 Å by cryo-electron microscopy. PT-AROD adopts an octameric structure with D4 symmetry, which is consistent with its molecular weight in solution, estimated by mass photometry. Comparative analysis of this structure with that determined using X-ray crystallography reveals open and closed forms of the lid-like loop at the entrance to the substrate pocket. Furthermore, mutation of Glu493, located at the substrate binding site, diminishes substrate selectivity, suggesting that this residue contributes significantly to the high selectivity of PT-AROD.

通过冷冻电镜和 X 射线晶体学研究 L-精氨酸氧化酶的开放和封闭结构。
L- 精氨酸氧化酶(AROD,EC 1.4.3.25)是一种以黄素腺嘌呤二核苷酸(FAD)为辅助因子催化 L- 精氨酸脱氨的氧化还原酶。最近从假单胞菌 TPU 7192(PT-AROD)中发现的 AROD 对 L-精氨酸具有高选择性。这种酶可用于精确检测生物样本中的 L-精氨酸。然而,依赖于 FAD 的 AROD 的结构特征仍然未知。在此,我们通过低温电子显微镜以 2.3 Å 的分辨率报告了 PT-AROD 的结构。PT-AROD 采用 D4 对称的八分子结构,这与其在溶液中的分子量(通过质量光度法估算)相符。将该结构与利用 X 射线晶体学确定的结构进行比较分析,可以发现底物口袋入口处的盖状环路有开放和封闭两种形式。此外,位于底物结合部位的 Glu493 发生突变后,底物选择性降低,这表明该残基对 PT-AROD 的高选择性起了重要作用。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
Journal of biochemistry
Journal of biochemistry 生物-生化与分子生物学
CiteScore
4.80
自引率
3.70%
发文量
101
审稿时长
4-8 weeks
期刊介绍: The Journal of Biochemistry founded in 1922 publishes the results of original research in the fields of Biochemistry, Molecular Biology, Cell, and Biotechnology written in English in the form of Regular Papers or Rapid Communications. A Rapid Communication is not a preliminary note, but it is, though brief, a complete and final publication. The materials described in Rapid Communications should not be included in a later paper. The Journal also publishes short reviews (JB Review) and papers solicited by the Editorial Board.
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