2-pyrone-4,6-dicarboxylate

• 分子结构分类: 有机化合物 - 有机杂环化合物 - 吡喃
• 英文名称: 2-pyrone-4,6-dicarboxylate
• 英文别名: 2-oxo-2H-pyran-4,6-dicarboxylate；6-oxopyran-2,4-dicarboxylate
• 化学式: C₇H₂O₆
• 分子量: 182.089
• InChiKey: VRMXCPVFSJVVCA-UHFFFAOYSA-L

计算性质

• 辛醇/水分配系数(LogP)：
  1.1
• 重原子数：
  13
• 可旋转键数：
  0
• 环数：
  1.0
• sp3杂化的碳原子比例：
  0.0
• 拓扑面积：
  107
• 氢给体数：
  0
• 氢受体数：
  6

反应信息

• 作为反应物：
  描述：

  2-pyrone-4,6-dicarboxylate 在 LigI hydrolase 、 LigJ hydratase 、 LigK aldolase 、 Novosphingobium sp. strain KA₁ LigU isomerase 、 还原型辅酶Ⅰ 、 malate dehydrogenase 作用下， 以 aq. phosphate buffer 、 重水 为溶剂， 生成 L-malate
  参考文献：
  名称：

  木质素细菌降解中催化烯丙基异构化的酶LigU的结构和化学反应机理。

  摘要：

  来自Novosphingobium sp。的LigU。菌株KA1催化（4 E）-草次戊酸（OMA）异构化为（3 Z）-2-酮--4-羧基-3-己烯二酸酯（KCH）作为原儿茶酸（PCA）4,5-裂解途径的一部分木质素的降解。通过X射线晶体学确定野生型酶的载脂蛋白形式的三维结构，并且在底物OMA存在下确定K66M突变体酶的结构。LigU是一种同源二聚体，不需要具有二氨基庚二酸酯差向异构酶结构折叠的辅因子或金属离子，它由具有相似拓扑结构的两个域组成。每个结构域具有一个中心α-螺旋，该α-螺旋被由反平行β链组成的β-桶围绕。活动站点位于两个域的缝隙中。1个H核磁共振波谱证明该酶在异构化反应中催化KCH的C5处的pro- S氢与D 2 O的交换。溶剂-氘交换实验表明，Lys-66的突变消除了C5处的同位素交换，而C100的突变取消了C3处的同位素交换。这两个残基在LigU活性位点中的定位与反应机制一

  DOI：

  10.1021/acs.biochem.9b00549
• 作为产物：
  描述：

  2-hydroxy-2H-pyran-4,6-dicarboxylate 、 NADP⁺ 生成 2-pyrone-4,6-dicarboxylate 、 氢(+1)阳离子 、 NADPH(4-)
  参考文献：
  名称：

  Genetic and Biochemical Characterization of 4-Carboxy-2-Hydroxymuconate-6-Semialdehyde Dehydrogenase and Its Role in the Protocatechuate 4,5-Cleavage Pathway in Sphingomonas paucimobilis SYK-6

  摘要：

  摘要 原儿茶酸盐（PCA）是鞘氨醇单胞菌（Sphingomonas paucimobilis）木质素降解途径中的关键中间代谢产物。 Sphingomonas paucimobilis SYK-6通过PCA 4,5-裂解途径代谢为丙酮酸和草酰乙酸。我们鉴定了 4-羧基-2-羟基琥珀酸-6-半乳糖醛（CHMS）脱氢酶基因 ( ligC ).CHMS是PCA的4,5-裂解产物，通过LigC转化为2-吡喃酮-4,6-二羧酸盐（PDC）。我们发现 位于 位于 编码大亚基的 编码 PCA 4,5-二氧合酶的大亚基。我们发现 基因 基因由一个 945 bp 的开放阅读框组成，编码一个分子质量为 34,590 Da 的多肽。推导的氨基酸序列为 氨基酸序列 的氨基酸序列与 3-氯苯甲酸酯 顺式 -二氢二醇脱氢酶有 19%至 20%的相同性。 Alcaligenes 菌株 BR60 和邻苯二甲酸酯 顺式 -二氢二醇脱氢酶的 假单胞菌 NMH102-2和 伯克霍尔德氏菌 DBO1，它们与第一、第二和第三类微生物醇脱氢酶无关（M. F. Reid 和 C. A. Fewson，Crit. Rev. Microbiol.）该 ligC 基因在 大肠杆菌中表达 并将 LigC 纯化至接近均一。在 NADP + 电喷雾质谱法和气相色谱质谱法证实了 LigC 催化 CHMS 产生 PDC。LigC 是一种同源二聚体。估计其等电点、最适 pH 值和最适温度分别为 5.3、8.0 和 25°C。其 K m 的 K m + 的 K m 估计为 24.6 ± 1.5 μM，比 NADP + (252 ± 3.9 μM)低约 10 倍。NAD K m 在 NADP + 和 NAD + 分别为 26.0 ± 0.5 和 20.6 ± 1.0 μM。破坏 ligC 的 S. paucimobilis SYK-6 阻止了香草酸盐的生长。在用香草酸盐与ligC-6 和ligC-6 的整个细胞孵育期间，只有 PCA 被积累。 ligC 插入突变体（DLC）的整个细胞中只积累了五氯苯甲醚，这表明 DLC 缺乏五氯苯甲醚-4,5-二氧酶活性。然而，引入 ligC 可恢复其在香草酸上的生长能力。PDC被认为是 ligAB 基因表达的诱导剂。

  DOI：

  10.1128/jb.182.23.6651-6658.2000

文献信息

• Purification and Properties of 2-Pyrone-4,6-Dicarboxylate Hydrolase
  作者：Kiyofumi MARUYAMA

  DOI：10.1093/oxfordjournals.jbchem.a134210

  日期：1983.2

  A hydrolase which catalyzes specifically the interconversion between 2-pyrone-4,6-dicarboxylate and 4-oxalmesaconate was purified about 410-fold with a 16% yield from cell-free extracts of Pseudomonas ochraceae grown with phthalate. Upon disc gel electrophoresis, the enzyme preparation gave a single band which was coincident with the enzyme activity. The molecular weight of the enzyme was estimated

  从邻苯二甲酸酯生长的无假单胞菌提取物中，水解催化特定水解2-吡喃酮4,6-二羧酸酯和4-草酸酯的水解酶约410倍，收率16％。经圆盘凝胶电泳后，酶制剂产生一条与酶活性一致的条带。通过在Sephadex G-75上进行凝胶过滤，估计该酶的分子量为31,000，通过十二烷基硫酸钠凝胶电泳估计该酶的分子量为33,000。通过等电聚焦测定该酶的等电点为pH 5.49。该酶对2-pyrone-4,6-dicarboxylate具有特异性，而其他各种内酯均不能用作底物。2-吡喃酮4-，6-二羧酸酯水解，4-草酸酯的形成和质子产生的化学计量约为1：1：1。水解和合成2-pyrone-4,6-dicarboxylate的最佳pH分别为8.5和6.0。2-pyrone-4,6-dicarboxylate和4-oxalmesaconate的Km值分别为87和26 microM。在pH 8.5下，处于平衡状态的
• 2-pyrone-4,6-dicarboxylic acid, a catabolite of gallic acids in Pseudomonas species
  作者：P J Kersten、S Dagley、J W Whittaker、D M Arciero、J D Lipscomb

  DOI：10.1128/jb.152.3.1154-1162.1982

  日期：1982.12

  2-Pyrone-4,6-dicarboxylate hydrolase was purified from 4-hydroxybenzoate-grown Pseudomonas testosteroni. Gel filtration and electrophoretic measurements indicated that the preparation was homogeneous and gave a molecular weight of 37,200 for the single subunit of the enzyme. Hydrolytic activity was dependent upon a functioning sulfhydryl group(s) and was freely reversible; the equilibrium position was dependent upon pH, with equimolar amounts of pyrone and open-chain form present at pH 7.9. Since the hydrolase was strongly induced when the nonfluorescent organisms P. testosteroni and P. acidovorans grew with 4-hydroxybenzoate, it is suggested that 2-pyrone-4,6-dicarboxylate is a normal intermediate in the meta fission degradative pathway of protocatechuate. Laboratory strains of fluorescent pseudomonads did not metabolize 2-pyrone-4,6-dicarboxylate, but a strain of P. putida was isolated from soil that utilized this compound for growth; the hydrolase was then induced, but it was absent from extracts of 4-hydroxybenzoate-grown cells that readily catabolized protocatechuate by ortho fission reactions. 2-Pyrone-4,6-dicarboxylic acid was the major product formed when gallic acid was oxidized by purified protocatechuate 3,4-dioxygenase. Protocatechuate 4,5-dioxygenase gave only the open-chain ring fission product when gallic acid was oxidized, but the enzyme attacked 3-O-methylgallic acid, giving 2-pyrone-4,6-dicarboxylic acid as the major product. Cell suspensions of 4-hydroxybenzoate-grown P. testosteroni readily oxidized 3-O-methylgallate with accumulation of methanol.

  2-吡喃-4,6-二羧酸水解酶从生长在4-羟基苯甲酸上的睾丸酮假单胞菌中纯化。凝胶过滤和电泳测量表明该制备是均一的，并给出了该酶单个亚基的分子量为37,200。水解活性取决于功能性巯基，并且是自由可逆的。平衡位置取决于pH，在pH 7.9时呈现出吡喃和开链形式的等摩尔量。由于当非荧光菌P. testosteroni和P. acidovorans在4-羟基苯甲酸上生长时，水解酶被强烈诱导，因此建议2-吡喃-4,6-二羧酸是原儿茶酸的代谢途径中间体。荧光假单胞菌的实验室菌株不代谢2-吡喃-4,6-二羧酸，但是从土壤中分离出一株利用该化合物生长的P. putida菌株，然后诱导水解酶，但在容易通过邻位裂解反应降解原儿茶酸的4-羟基苯甲酸生长的细胞提取物中缺少该酶。当用纯化的原儿茶酸3,4-双加氧酶氧化没食子酸时，2-吡喃-4,6-二羧酸是主要产物。当氧化没食子酸时，原儿茶酸4,5-双加氧酶只产生开链环裂解产物，但是该酶攻击3-O-甲基没食子酸，将2-吡喃-4,6-二羧酸作为主要产物。4-羟基苯甲酸生长的P. testosteroni细胞悬液容易氧化3-O-甲基没食子酸，并积累甲醇。
• Purification and Properties of α-Hydroxy-γ-Carboxymuconic ɛ-Semialdehyde Dehydrogenase
  作者：Kiyofumi MARUYAMA、Nakao ARIGA、Mitsushige TSUDA、Kazuo DEGUCHI

  DOI：10.1093/oxfordjournals.jbchem.a132002

  日期：1978.4

  α-Hydroxy-γ-carboxymuconic ɛ-semialdehyde (HCMS) dehydrogenase was purified about 180-fold with a yield of 22% from a cell-free extract of Pseudomonas ochraceae. The enzyme was adsorbed tightly on blue dextran, possibly at its coenzyme binding sites. This finding made possible a simple purification step by molecular exclusion chromatography on Sephadex G-200. The enzyme adsorbed on blue dextran was eluted in the void volume and was well separated from contaminating proteins which were eluted at various positions. The purified enzyme migrated as a single band on disc gel electrophoresis. The molecular weight of the enzyme as determined by gel filtration on Sephadex G-200 was 67,000. Polycrylamide gel electrophoresis of the enzyme after sodium dodecyl sulfate denaturation gave a single band at a position corresponding to a molecular weight of 35,000. The enzyme showed a simple protein absorption in the UV region and no significant absorption in the visible region. The enzyme activity was not affected by various metal ions, metal chelating reagents or reducing reagents, but was strongly inhibited by various sulfhydryl reagents. Both NAD and NADP were found to be efficient coenzymes. Various aldehyde derivatives such as acetaldehyde, propionaldehyde, n-butyraldehyde, iso-butyraldehyde, succinaldehyde, crotonaldehyde, chloral hydrate, glyoxylic acid, DL-glyceraldehyde, benzaldehyde, salicylaldehyde, glucose, and protocatechualdehyde did not serve as substrates. The stoichiometry of HCMS consumption and coenzyme reduction was approximately 1: 1. The reaction product, which appeared to be α-hydroxy-γ-carboxymuconic acid, showed an absorption maximum at 311 nm at pH 8.0.

  α-羟基-γ-羧基-戊二酸半醛（HCMS）脱氢酶从黄曲假单胞菌的无细胞提取物中纯化约180倍，得率为22%。该酶紧密吸附在蓝色葡聚糖上，可能吸附在其辅酶结合位点上。这一发现使得通过Sephadex G-200分子排阻色谱法进行简单纯化成为可能。吸附在蓝色葡聚糖上的酶在空隙体积中洗脱，并与在不同位置洗脱的污染蛋白很好地分离。纯化的酶在圆盘凝胶电泳中作为单个条带迁移。通过Sephadex G-200凝胶过滤法测定的酶分子量为67,000。十二烷基硫酸钠变性后，酶的多聚酰胺凝胶电泳在对应分子量为35,000的位置出现单个条带。该酶在紫外区显示简单的蛋白质吸收，在可见区没有明显的吸收。酶活性不受各种金属离子、金属螯合试剂或还原试剂的影响，但受到各种巯基试剂的强烈抑制。NAD和NADP都被发现是有效的辅酶。各种醛类衍生物，如乙醛、丙醛、正丁醛、异丁醛、琥珀醛、巴豆醛、水合氯醛、乙醛酸、DL-甘油醛、
• Interaction of 4-Carboxy-2-Hydroxymuconate-6-Semialdehyde Dehydrogenase with Reactive Blue 2 and Related Dyes
  作者：Kiyofumi Maruyama

  DOI：10.1093/oxfordjournals.jbchem.a122334

  日期：1988.4

  Steady-state kinetic analyses suggest that Pseudomonas ochraceae 4-carboxy-2-hydroxymuconate-6-semialdehyde dehydrogenase (4-carboxy-2-hydroxy-cis, cis-muconate-6-semialdehyde: NADP+ oxidoreductase [EC 1.2.1.45]) functions through an ordered BiBi mechanism. The enzyme binds one NADP+ molecule per subunit with a Kd of 4.8±0.8 μM. The enzyme is adsorbed to a Blue Sepharose CL-6B column and can be eluted therefrom with reagents having high affinity for the enzyme such as NADP+, NAD+, ATP, and Reactive Blue 2. Equilibrium dialysis and difference spectral titration show the binding of four molecules of Reactive Blue 2 per enzyme subunit. Two of these dye molecules show high-affinity binding with a Kd of 0.03±0.02 μM. The resulting 1: 2 enzyme-dye complex can be isolated by gel filtration on Bio-Gel P-6. The kinetic, spectroscopic, and chromatographic properties of the complex indicate that the dye-binding sites are different from the coenzyme binding site. The other two dye molecules, in contrast, bind loosely with a Kd of 0.8±0.5 μM to a site overlapping the coenzyme binding site. This is confirmed by the following findings: NADP+ effectively abolishes the difference spectrum associated with the enzyme-dye binding, and the slope of the double reciprocal plot showing the competitive inhibition of the dye (K1 =0.20±0.02 μM) with respect to NADP+ linearly depends on the square of the dye concentration. Essentially similar results are also obtained with methoxy Reactive Blue 2 and Reactive Blue 4. The binding of Blue Dextran-2000 to the enzyme is somewhat unlike that of Reactive Blue 2, possibly due to insufficient contact of the dye chromophore with the enzyme.

  稳态动力学分析表明，黄曲假单胞菌4-羧基-2-羟基-顺式-穆孔酸-6-半醛脱氢酶（4-羧基-2-羟基-顺式-穆孔酸-6-半醛：NADP+氧化还原酶[EC 1.2.1.45]）通过有序的双分子机制发挥作用。该酶每个亚基结合一个NADP+分子，结合常数（Kd）为4.8±0.8 μM。该酶吸附在Blue Sepharose CL-6B柱上，可用对酶具有高亲和力的试剂（如NADP+、NAD+、ATP和活性蓝2）从柱上洗脱。平衡透析和差分光谱滴定显示，每个酶亚基结合四个活性蓝2分子。其中两个染料分子表现出高亲和力结合，结合常数（Kd）为0.03±0.02 μM。由此形成的1:2酶-染料复合物可通过Bio-Gel P-6凝胶过滤法分离。该复合物的动力学、光谱和色谱性质表明，染料结合位点与辅酶结合位点不同。相比之下，另外两个染料分子与辅酶结合位点重叠的位点结合松散，结合常数（Kd）为0.8±
• Isolation and Identification of the Reaction Product of α-Hydroxy-γ-Carboxymuconic ε-Semialdehyde Dehydrogenase 1
  作者：Kiyofumi MARUYAMA

  DOI：10.1093/oxfordjournals.jbchem.a132687

  日期：1979.12

  The reaction product of enzymic dehydrogenation of alpha-hydroxy-gamma-carboxymuconic epsilon-semialdehyde (HCMS) was isolated. The analytical data (elemental analyses, IR spectra, mass spectra, proton NMR spectra, and UV spectra) showed that the product was not alpha-hydroxy-gamma-carboxymuconic acid (HCMA), the expected primary product of HCMS dehydrogenation, but a lactone of HCMA. The structure

  分离出α-羟基-γ-羧基粘康酸ε-半醛（HCMS）的酶促脱氢反应产物。分析数据（元素分析，红外光谱，质谱，质子NMR光谱和UV光谱）表明，该产物不是α-羟基-γ-羧基粘康酸（HCMA），它是HCMS脱氢的预期主要产物，而是内酯HCMA。内酯的结构初步确定为α-吡喃酮4，6-二羧酸。通过纯化的NAD（P）连接的HCMS脱氢酶将HCMS化学计量地转化为内酯。内酯可通过在邻苯二甲酸上生长的无extract提取物（由se菜假单胞菌制备）进行积极代谢，这表明它可能是原儿茶酸细菌代谢的代谢中间体。此外，