D-aspartate

• 分子结构分类: 有机化合物 - 有机酸及其衍生物 - 羧酸及其衍生物
• 英文名称: D-aspartate
• 英文别名: L-Iso-Aspartate；(2R)-2-amino-4-hydroxy-4-oxobutanoate
• 化学式: C₄H₆NO₄
• 分子量: 132.096
• InChiKey: CKLJMWTZIZZHCS-UWTATZPHSA-M

计算性质

• 辛醇/水分配系数(LogP)：
  -2.6
• 重原子数：
  9
• 可旋转键数：
  1
• 环数：
  0.0
• sp3杂化的碳原子比例：
  0.5
• 拓扑面积：
  108
• 氢给体数：
  1
• 氢受体数：
  4

反应信息

• 作为反应物：
  描述：

  trans-{Co((1R,2R)-N,N'-disalicylidene-1,2-cyclohexanediamine)(H2O)2} 、 D-aspartate 以 甲醇 、 水 为溶剂， 生成 trans-(Co((1R,2R)-N,N'-disalicylidene-1,2-cyclohexanediamine)(CH3COO)(H2O)) 、 Δ-β2-{Co((1R,2R)-N,N'-disalicylidene-1,2-cyclohexanediamine)(L-asp)}
  参考文献：
  名称：

  The Stereochemistry and Reactivity of Metal-Schiff Base Complexes. VII. Contribution of Hydrophobic Interligand Interaction to Chiral Recognition of Phenylalaninate and Tryptophanate with (1R,2R)-N,N′-Disalicylidene-1,2-cyclohexanediaminecobalt(III) Complex

  摘要：

  Δ-β2-型混配体钴(III)配合物的稳定常数K1是通过光谱法在22°C下，在含有乙酸盐缓冲液(0.3 mol dm−3)的水-甲醇(2:3体积比)混合溶液中测定的。这些配合物由手性四齿席夫碱(sal-(R,R)-chxn)和d-或l-氨基酸根(aa− = gly、ala、val、leu、thr、phe、trp、pro、asp、asn和glu)组成。sal-(R,R)-chxn是由水杨醛和(R,R)-1,2-环己二胺衍生而来。 K1值范围在5.6×106到1.2×109 mol−1 dm3之间，除了d-phe、d-trp、d-asp、d-asn和d-pro外，都符合线性自由能关系。对于d-phe、d-trp、d-asp和d-asn，它们的稳定常数是相应l-氨基酸的5-30倍。 基于构象分析和1H NMR光谱，文章讨论了d-phe和d-trp配合物的特殊稳定性，认为这与席夫碱配体和氨基酸侧链之间的芳香环层间堆积有关。

  DOI：

  10.1246/bcsj.63.138
• 作为产物：
  描述：

  aspartate anion 生成 D-aspartate
  参考文献：
  名称：

  The mcyF gene of the microcystin biosynthetic gene cluster from Microcystis aeruginosa encodes an aspartate racemase

  摘要：

  微囊藻毒素是一种具有肝毒性的非核糖体肽，由多个淡水蓝藻属产生。在其他酶活性（尤其是肽合成酶和聚酮合成酶的活性）中，微囊藻毒素的生物合成需要外消旋酶提供 d-天门冬氨酸和 d-谷氨酸。在此，我们报告了一个开放阅读框 mcyF 的克隆、表达和表征，它是铜绿微囊藻菌株 PCC 7806 中参与微囊藻毒素生物合成的 mcy 基因簇的一部分。保守的氨基酸序列基序表明，McyF 蛋白具有天冬氨酸外消旋酶的功能。在单细胞蓝藻 Synechocystis PCC 6803 中异源表达 mcyF，可获得具有活性的 His6 标记蛋白，该蛋白可通过 Ni2+-Nitriloacetate 亲和色谱法纯化至均一。纯化的重组蛋白以不依赖于吡哆醛-5′-磷酸的方式消旋天门冬氨酸，但不消旋谷氨酸。此外，我们还在铜绿微囊藻 PCC 7806 基因组中发现了一个位于 mcy 基因簇之外的推定谷氨酸消旋酶基因。该谷氨酸消旋酶基因的同源物存在于所有被检测的微囊藻菌株中，而 mcyF 则只能在产生微囊藻毒素的菌株中检测到。

  DOI：

  10.1042/bj20030396

文献信息

• Purification and Characterization of Novel<i>N</i>-Acyl-<scp>D</scp>-aspartate Amidohydrolase from<i>Alcaligenes xylosoxydans</i>subsp.<i>xylosoxydans</i>A-6
  作者：Mitsuaki Moriguchi、Kenji Sakai、Yutaka Katsuno、Tetsuyoshi Maki、Mamoru Wakayama

  DOI：10.1271/bbb.57.1145

  日期：1993.1

  Alcaligenes xylosoxydans subsp. xylosoxydans A-6 (Alcaligenes A-6) produced N-acyl-D-aspartate amidohydrolase (D-AAase) in the presence of N-acetyl-D-aspartate as an inducer. The enzyme was purified to homogeneity. The enzyme had a molecular mass of 56 kDa and was shown by sodium dodecyl sulfate (SDS)–polyacrylamide gel electrophoresis (PAGE) to be a monomer. The isoelectric point was 4.8. The enzyme had maximal activity at pH 7.5 to 8.0 and 50°C, and was stable at pH 8.0 and up to 45°C. N-Formyl (Km=12.5 mM), N-acetyl (Km=2.52 mM), N-propionyl (Km=0.194 mM), N-butyryl (Km=0.033 mM), and N-glycyl (Km =1.11 mM) derivatives of D-aspartate were hydrolyzed, but N-carbobenzoyl-D-aspartate, N-acetyl-L-aspartate, and N-acetyl-D-glutamate were not substrates. The enzyme was inhibited by both divalent cations (Hg2+, Ni2+, Cu2+) and thiol reagents (N-ethylmaleimide, iodoacetic acid, dithiothreitol, and p-chloromercuribenzoic acid). The N-terminal amino acid sequence and amino acid composition were analyzed.

  Alcaligenes xylosoxydans subsp. xylosoxydans A-6（Alcaligenes A-6）在以 N-乙酰-D-天冬氨酸为诱导剂的情况下产生 N-乙酰-D-天冬氨酸酰胺水解酶（D-AAase）。该酶已纯化至均一。十二烷基硫酸钠（SDS）-聚丙烯酰胺凝胶电泳（PAGE）显示，该酶的分子质量为 56 kDa，为单体。等电点为 4.8。该酶在 pH 值为 7.5 至 8.0 和 50°C 时活性最大，在 pH 值为 8.0 和高达 45°C 时稳定。可水解 D-天冬氨酸的 N-甲酰基（Km=12.5 mM）、N-乙酰基（Km=2.52 mM）、N-丙酰基（Km=0.194 mM）、N-丁酰基（Km=0.033 mM）和 N-甘氨酰（Km=1.11 mM）衍生物，但 N-羧基苯甲酰基-D-天冬氨酸、N-乙酰基-L-天冬氨酸和 N-乙酰基-D-谷氨酸不是底物。该酶受到二价阳离子（Hg2+、Ni2+、Cu2+）和硫醇试剂（N-乙基马来酰亚胺、碘乙酸、二硫苏糖醇和对氯巯基苯甲酸）的抑制。分析了 N 端氨基酸序列和氨基酸组成。
• Molecular identification of monomeric aspartate racemase from Bifidobacterium bifidum
  作者：Tatsuyuki Yamashita、Makoto Ashiuchi、Kouhei Ohnishi、Shin'ichiro Kato、Shinji Nagata、Haruo Misono

  DOI：10.1111/j.1432-1033.2004.04445.x

  日期：2004.12

  Bifidobacterium bifidum is a useful probiotic agent exhibiting health‐promoting properties and contains d‐aspartate as an essential component of the cross‐linker moiety in the peptidoglycan. To help understand d‐aspartate biosynthesis in B. bifidum NBRC 14252, aspartate racemase, which catalyzes the racemization of d‐ and l‐aspartate, was purified to homogeneity and characterized. The enzyme was a monomer with a molecular mass of 27 kDa. This is the first report showing the presence of a monomeric aspartate racemase. Its enzymologic properties, such as its lack of cofactor requirement and susceptibility to thiol‐modifying reagents in catalysis, were similar to those of the dimeric aspartate racemase from Streptococcus thermophilus. The monomeric enzyme, however, showed a novel characteristic, namely, that its thermal stability significantly increased in the presence of aspartate, especially the d‐enantiomer. The gene encoding the monomeric aspartate racemase was cloned and overexpressed in Escherichia coli cells. The nucleotide sequence of the aspartate racemase gene encoded a peptide containing 241 amino acids with a calculated molecular mass of 26 784 Da. The recombinant enzyme was purified to homogeneity and its properties were almost the same as those of the B. bifidum enzyme.

  *Bifidobacterium bifidum* 是一种有益的益生菌，具有促进健康的特性，并且其细胞壁肽聚糖中包含D-天冬氨酸作为交联单元的重要组成部分。为了更好地理解NBRC 14252株*B. bifidum* 中D-天冬氨酸的生物合成机制，研究人员纯化并表征了天冬氨酸消旋酶，该酶催化D-天冬氨酸和L-天冬氨酸之间的消旋作用。该酶为单体蛋白，分子量为27千道尔顿。这是首次报道单体天冬氨酸消旋酶的存在。其生化特性，例如不需要辅助因子以及易受含硫修饰试剂的影响，与来源于*Streptococcus thermophilus* 的二聚体天冬氨酸消旋酶相似。然而，单体酶表现出一种新型特性，即其热稳定性在天冬氨酸存在下显著增加，尤其是D-天冬氨酸。编码单体天冬氨酸消旋酶的基因已被克隆并在*Escherichia coli* 细胞中过表达。该酶基因的核苷酸序列编码了一个由241个氨基酸组成的多肽，理论计算分子量为26,784道尔顿。重组酶经纯化后表现出与*B. bifidum* 原生酶几乎相同的酶学特性。
• A diverse superfamily of enzymes with ATP-dependent carboxylate-amine/thiol ligase activity
  作者：Michael Y. Galperin、Eugene V. Koonin

  DOI：10.1002/pro.5560061218

  日期：——

  tubulin-tyrosine ligase, and three enzymes of purine biosynthesis. All these enzymes possess ATP-dependent carboxylate-amine ligase activity, and their catalytic mechanisms are likely to include acylphosphate intermediates. The ATP-grasp superfamily also includes succinate-CoA ligase (both ADP-forming and GDP-forming variants), malate-CoA ligase, and ATP-citrate lyase, enzymes with a carboxylate-thiol ligase activity

  最近开发的用于序列数据库搜索的PSI-BLAST方法和用于基序分析的方法用于定义和扩展具有异常核苷酸结合折叠（称为棕榈酸酯或ATP抓握折叠）的酶超家族。除了具有已知三维结构的D-丙氨酸-D-丙氨酸连接酶，谷胱甘肽合成酶，生物素羧化酶和氨基甲酰磷酸合成酶外，还在核糖体蛋白S6修饰酶（RimK），尿素中预测到ATP抓图结构域酰胺酶，微管蛋白酪氨酸连接酶和嘌呤生物合成的三种酶。所有这些酶都具有ATP依赖的羧酸胺连接酶活性，其催化机制可能包括酰基磷酸酯中间体。ATP抓取超家族还包括琥珀酸CoA连接酶（形成ADP的变体和形成GDP的变体），苹果酸-CoA连接酶和ATP-柠檬酸裂合酶，具有羧酸盐-硫醇连接酶活性的酶以及一些未鉴定的蛋白质。这些发现显着扩展了ATP抓握酶底物的种类以及它们所涉及的生化途径的范围，并证明了结构比较和强大的序列分析方法之间的互补性。
• Aslfm, the D-Aspartate Ligase Responsible for the Addition of D-Aspartic Acid onto the Peptidoglycan Precursor of Enterococcus faecium
  作者：Samuel Bellais、Michel Arthur、Lionnel Dubost、Jean-Emmanuel Hugonnet、Laurent Gutmann、Jean van Heijenoort、Raymond Legrand、Jean-Paul Brouard、Louis Rice、Jean-Luc Mainardi

  DOI：10.1074/jbc.m600114200

  日期：2006.4

  protein superfamily, which includes a diverse set of enzymes catalyzing ATP-dependent carboxylate-amine ligation reactions. Aslfm specifically ligated the beta-carboxylate of D-Asp to the epsilon-amino group of L-Lys in the nucleotide precursor UDP-N-acetylmuramyl-pentapeptide. D-iso-asparagine was not a substrate of Aslfm, indicating that the presence of this amino acid in the peptidoglycan of E. faecium

  D-天冬氨酸连接酶一直是革兰氏阳性细菌的肽聚糖组装途径中最后一个未确定的肽键形成酶。在这里，我们显示粪肠球菌的两个基因簇编码天冬氨酸消旋酶（Racfm）和连接酶（Aslfm），以将D-Asp掺入肽聚糖前体的侧链。Aslfm被确定为ATP-抓蛋白超家族的新成员，该家族包括催化ATP依赖的羧酸盐-胺连接反应的多种酶。Aslfm将D-Asp的β-羧酸盐特异地连接到核苷酸前体UDP-N-乙酰基村mura基五肽中L-Lys的ε-氨基上。D-异天冬酰胺不是Aslfm的底物，表明该氨基酸存在于E.的肽聚糖中。粪便是D-Asp加入前体后α-羧基酰胺化的结果。粪肠球菌中编码Racfm和Aslfm的基因的异源表达导致产生被D-Asp代替L-Ala2取代的干肽，为这些酶的体内特异性和功能提供了证据。令人惊讶的是，对跨桥的测序表明，受体和供体底物中青霉素结合蛋白的d，d-转肽酶活性可耐受D-Asp取代L-Ala2
• Functional and Structural Characterization of D-Aspartate Oxidase from Porcine Kidney: Non-Michaelis Kinetics due to Substrate Activation
  作者：A. Yamamoto、H. Tanaka、T. Ishida、K. Horiike

  DOI：10.1093/jb/mvm041

  日期：——

  d-Aspartate oxidase (DDO, EC 1.4.3.1) catalyzes dehydrogenation of d-aspartate to iminoaspartate and the subsequent re-oxidation of reduced FAD with O2 to produce hydrogen peroxide. In the mammalian neuroendocrine system, d-aspartate, a natural substrate, plays important roles in the regulation of the synthesis and secretion of hormones. To elucidate the kinetic and structural properties of native DDO, we purified DDO from porcine kidney to homogeneity, cloned the cDNA, and overexpressed the enzyme in Escherichia coli. The purified DDO was a homotetramer with tightly-bound FAD. The enzyme consisted of 341 amino acids and had GAGVMG as the dinucleotide binding motif and a C-terminal SKL peroxisomal-targeting signal sequence. Porcine DDO showed a strong affinity for meso-tartrate (Kd = 118 μM). The oxidase exhibited pronounced substrate activation at d-aspartate and d-glutamate concentrations, [S], higher than 0.2 and 4 mM, respectively, and the [S]/v versus [S] plot showed marked downward curvature (v, the initial velocity), whereas substrate inhibition occurred with N-methyl-d-aspartate. These kinetic properties of DDO suggested that at high substrate concentrations, the FAD-reduced form of the enzyme also catalyzes the reaction: the oxidative half-reaction precedes the reductive one. The present direct approach to the analysis of non-Michaelis kinetics is indispensable for understanding the functional properties of DDO.

  d-天门冬氨酸氧化酶（DDO，EC 1.4.3.1）催化 d-天门冬氨酸脱氢为亚氨基天门冬氨酸，随后还原的 FAD 与 O2 再氧化产生过氧化氢。在哺乳动物的神经内分泌系统中，d-天门冬氨酸是一种天然底物，在调节激素的合成和分泌方面发挥着重要作用。为了阐明原生 DDO 的动力学和结构特性，我们从猪肾中纯化出均一的 DDO，克隆了 cDNA，并在大肠杆菌中过表达了该酶。纯化的 DDO 是一种同源四聚体，与 FAD 紧密结合。该酶由 341 个氨基酸组成，二核苷酸结合基序为 GAGVMG，C 端为 SKL 过氧化物酶体靶向信号序列。猪 DDO 对中酒石酸有很强的亲和力（Kd = 118 μM）。当 d-天冬氨酸和 d-谷氨酸浓度[S]分别高于 0.2 mM 和 4 mM 时，该氧化酶表现出明显的底物活化，[S]/v 与[S]的关系图显示出明显的向下弯曲（v，初始速度），而 N-甲基-d-天冬氨酸则出现底物抑制。DDO 的这些动力学特性表明，在底物浓度较高时，该酶的 FAD 还原型也会催化反应：氧化半反应先于还原半反应。目前直接分析非迈克尔动力学的方法对于了解 DDO 的功能特性是不可或缺的。