Structure of the flavocoenzyme of two homologous amine oxidases: monomeric sarcosine oxidase and N-methyltryptophan oxidase.
Structure of the flavocoenzyme of two homologous amine oxidases: monomeric sarcosine oxidase and N-methyltryptophan oxidase.
复制标题
两种同源胺氧化酶的黄素辅酶的结构：单体肌氨酸氧化酶和 N-甲基色氨酸氧化酶。
DOI：
10.1021/bi982955o
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发表时间：
1999
期刊：
影响因子：
--
通讯作者：
Jorns,MS
中科院分区：
文献类型：
--
作者：
Wagner,MA;Khanna,P;Jorns,MS
单体肌氨酸氧化酶（MSOX）和N-甲基色氨酸氧化酶（MTOX）是同源酶，分别催化肌氨酸（N-甲基甘氨酸）和N-甲基-L-色氨酸的氧化去甲基化。 MSOX 在各种细菌在肌氨酸上生长时被诱导。 MTOX 是 E。代谢功能未知的大肠杆菌酶。两种酶都含有共价结合的黄素。通过电喷雾质谱法判断，共价黄素处于FAD水平。这些数据首次证明 MTOX 是一种黄素蛋白。以下观察结果表明 8α-(S-半胱氨酰)FAD 是芽孢杆菌属 MSOX 中的共价黄素。 B-0618和MTOX。含有 FMN 的肽是通过用胰蛋白酶、胰凝乳蛋白酶和磷酸二酯酶消化 MSOX 或 MTOX 制备的，具有 8α-(S-半胱氨酰)黄素的吸收和荧光特性，并且可以与脱辅基黄素氧还蛋白结合。通过特征吸光度变化和黄素荧光增加来判断，含 FMN 的肽中的硫醚键通过过甲酸氧化转化为砜。在用连二亚硫酸盐处理后，砜经历了预期的还原裂解反应，释放出未修饰的 FMN。 Cys315 被鉴定为 B 的 MSOX 中的共价 FAD 附着位点。 sp。 B-0618，根据含黄素胰蛋白酶肽 (GAVCMYT) 获得的序列判断。 Cys315 与来自其他细菌、MTOX (Cys308) 和哌可酸氧化酶（一种已知含有共价结合黄素的同源哺乳动物酶）的 MSOX 中的保守半胱氨酸对齐。这些酶中仅发现一个保守的半胱氨酸，表明 Cys308 是 MTOX 中的共价黄素附着位点。
Monomeric sarcosine oxidase (MSOX) andN-methyltryptophan oxidase (MTOX) are homologous enzymes that catalyze the oxidative demethylation of sarcosine (N-methylglycine) andN-methyl-l-tryptophan, respectively. MSOX is induced in various bacteria upon growth on sarcosine. MTOX is anE. colienzyme of unknown metabolic function. Both enzymes contain covalently bound flavin. The covalent flavin is at the FAD level as judged by electrospray mass spectrometry. The data provide the first evidence that MTOX is a flavoprotein. The following observations indicate that 8α-(S-cysteinyl)FAD is the covalent flavin in MSOX fromBacillus sp. B-0618and MTOX. FMN-containing peptides, prepared by digestion of MSOX or MTOX with trypsin, chymotrypsin, and phosphodiesterase, exhibited absorption and fluorescence properties characteristic of an 8α-(S-cysteinyl)flavin and could be bound to apo-flavodoxin. The thioether link in the FMN-containing peptides was converted to the sulfone by performic acid oxidation, as judged by characteristic absorbance changes and an increase in flavin fluorescence. The sulfone underwent a predicted reductive cleavage reaction upon treatment with dithionite, releasing unmodified FMN. Cys315 was identified as the covalent FAD attachment site in MSOX fromB. sp. B-0618, as judged by the sequence obtained for a flavin-containing tryptic peptide (GAVCMYT). Cys315 aligns with a conserved cysteine in MSOX from other bacteria, MTOX (Cys308) and pipecolate oxidase, a homologous mammalian enzyme known to contain covalently bound flavin. There is only one conserved cysteine found among these enzymes, suggesting that Cys308 is the covalent flavin attachment site in MTOX.