title: Crystal structure of a membrane-bound l-amino acid deaminase from Proteus vulgaris.
abstract: l-amino acid oxidases/deaminases (LAAOs/LAADs) are a class of oxidoreductases catalyzing the oxidative deamination of l-amino acids to α-keto acids. They are widely distributed in eukaryotic and prokaryotic organisms, and exhibit diverse substrate specificity, post-translational modifications and cellular localization. While LAAOs isolated from snake venom have been extensively characterized, the structures and functions of LAAOs from other species are largely unknown. Here, we reported crystal structure of a bacterial membrane-bound LAAD from Proteus vulgaris (pvLAAD) in complex with flavin adenine dinucleotide (FAD). We found that the overall fold of pvLAAD does not resemble typical LAAOs. Instead it, is similar to d-amino acid oxidases (DAAOs) with an additional hydrophobic insertion module on protein surface. Structural analysis and liposome-binding assays suggested that the hydrophobic module serves as an extra membrane-binding site for LAADs. Bacteria from genera Proteus and Providencia were found to encode two classes of membrane-bound LAADs. Based on our structure, the key roles of residues Q278 and L317 in substrate selectivity were proposed and biochemically analyzed. While LAADs on the membrane were proposed to transfer electrons to respiratory chain for FAD re-oxidization, we observed that the purified pvLAAD could generate a significant amount of hydrogen peroxide in vitro, suggesting it could use dioxygen to directly re-oxidize FADH2 as what typical LAAOs usually do. These findings provide a novel insights for a better understanding this class of enzymes and will help developing biocatalysts for industrial applications.
authors.affiliation: Research Center for Structural Biology and Drug Discovery, School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou 510006, China.
authors.forename: Yingchen
authors.initials: Y
authors.lastname: Ju
authors.affiliation: Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science and Technology of China, 96 Jinzhai Road, Hefei, Anhui 230027, China; Key Laboratory of Structural Biology, Chinese Academy of Sciences, 96 Jinzhai Road, Hefei, Anhui 230027, China.
authors.forename: Shuilong
authors.initials: S
authors.lastname: Tong
authors.affiliation: Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science and Technology of China, 96 Jinzhai Road, Hefei, Anhui 230027, China.
authors.forename: Yongxiang
authors.initials: Y
authors.lastname: Gao
authors.affiliation: Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science and Technology of China, 96 Jinzhai Road, Hefei, Anhui 230027, China; Key Laboratory of Structural Biology, Chinese Academy of Sciences, 96 Jinzhai Road, Hefei, Anhui 230027, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhao
authors.affiliation: Research Center for Structural Biology and Drug Discovery, School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou 510006, China.
authors.forename: Qi
authors.initials: Q
authors.lastname: Liu
authors.affiliation: Research Center for Structural Biology and Drug Discovery, School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou 510006, China.
authors.forename: Qiong
authors.initials: Q
authors.lastname: Gu
authors.affiliation: Research Center for Structural Biology and Drug Discovery, School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou 510006, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Xu
authors.affiliation: Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science and Technology of China, 96 Jinzhai Road, Hefei, Anhui 230027, China; Key Laboratory of Structural Biology, Chinese Academy of Sciences, 96 Jinzhai Road, Hefei, Anhui 230027, China. Electronic address: lwniu@ustc.edu.cn.
authors.forename: Liwen
authors.initials: L
authors.lastname: Niu
authors.affiliation: Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science and Technology of China, 96 Jinzhai Road, Hefei, Anhui 230027, China; Key Laboratory of Structural Biology, Chinese Academy of Sciences, 96 Jinzhai Road, Hefei, Anhui 230027, China. Electronic address: mkteng@ustc.edu.cn.
authors.forename: Maikun
authors.initials: M
authors.lastname: Teng
authors.affiliation: Research Center for Structural Biology and Drug Discovery, School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou 510006, China. Electronic address: zhuihao@mail.sysu.edu.cn.
authors.forename: Huihao
authors.initials: H
authors.lastname: Zhou
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry*; D055162:Biocatalysis; D020134:Catalytic Domain; D002462:Cell Membrane / Q000201:enzymology; D018360:Crystallography, X-Ray; D006861:Hydrogen Peroxide / Q000737:chemistry; D057927:Hydrophobic and Hydrophilic Interactions; D050830:L-Amino Acid Oxidase / Q000737:chemistry*; D008958:Models, Molecular; D011485:Protein Binding; D000072756:Protein Conformation, alpha-Helical; D011514:Proteus vulgaris / Q000201:enzymology*
chemical_list: D001426:Bacterial Proteins; D006861:Hydrogen Peroxide; D050830:L-Amino Acid Oxidase
keywords: Crystal structure; Deaminase; Insertion module; Membrane binding; Peroxide; l-amino acid oxidase