Saeki Y, Nozaki M, Senoh S
J Biol Chem. 1980 Sep 25;255(18):8465-71.
Both intradiol and proximal extradiol dioxygenases are thought to produce the same product, alpha-hydroxymuconic acid, when pyrogallol (3-hydroxycatechol) is used as a substrate. However, when these enzymes were reacted with pyrogallol, they gave different products. A proximal extradiol dioxygenase, metapyrocatechase (catechol:oxygen 2,3-d-oxidoreductase (decyclizing), EC 1.13.11.2), gave a product having an absorption maximum at 290 nm, which was gradually converted to a more stable compound having an absorption maximum at 239 nm. On the other hand, an intradiol dioxygenase, protocatechuate 3,4-dioxygenase (protocatechuate:oxygen 3,4-oxidoreductase (decyclizing), EC 1.13.11.3), gave a product having an absorption maximum at 300 nm. Based on the spectral data and direct comparison with authentic samples, the primary products obtained by the action of the former and the latter enzymes were identified as alpha-hydroxymuconic acid and 2-pyrone-6-carboxylic acid, respectively. While another intradiol dioxygenase, pyrocatechase (catechol:oxygen 1,2-oxidoreductase (decyclizing), EC 1.13.11.1), gave a mixture of nearly equimolar amounts of these two compounds. Isotope labeling experiments indicated that 1 atom of oxygen was incorporated in 2-pyrone-6-carboxylic acid from the atmosphere. Based on these findings, the reaction mechanism for the formation of 2-pyrone-6-carboxylic acid is discussed. This may be the first experimental evidence indicating the presence of a seven-membered lactone intermediate during the oxygenative cleavage of catechols, proposed by Hamilton (Hamilton, G.A. (1974) in Molecular Mechanisms of Oxygen Activation (Hayaishi, O., ed) pp. 405-451, Academic Press, New York).
当使用焦性没食子酸（3-羟基邻苯二酚）作为底物时，二醇内加氧酶和近端二醇外切加氧酶都被认为会产生相同的产物α-羟基粘康酸。然而，当这些酶与焦性没食子酸反应时，它们产生了不同的产物。一种近端二醇外切加氧酶，间位焦儿茶酚酶（儿茶酚：氧2,3-d-氧化还原酶（环化），EC 1.13.11.2），产生了一种在290nm处具有最大吸收的产物，该产物逐渐转化为一种在239nm处具有最大吸收的更稳定的化合物。另一方面，一种二醇内加氧酶，原儿茶酸3,4-双加氧酶（原儿茶酸：氧3,4-氧化还原酶（环化），EC 1.13.11.3），产生了一种在300nm处具有最大吸收的产物。基于光谱数据并与真实样品进行直接比较，前者和后者酶作用获得的主要产物分别被鉴定为α-羟基粘康酸和2-吡喃酮-6-羧酸。而另一种二醇内加氧酶，焦儿茶酚酶（儿茶酚：氧1,2-氧化还原酶（环化）EC 1.13.11.1）产生了这两种化合物近等摩尔量的混合物。同位素标记实验表明，大气中的1个氧原子被结合到2-吡喃酮-6-羧酸中。基于这些发现，讨论了2-吡喃酮-6-羧酸形成的反应机制。这可能是第一个实验证据，表明在儿茶酚的氧化裂解过程中存在七元内酯中间体，这是由汉密尔顿提出的（汉密尔顿，G.A.（1974年），《氧激活的分子机制》（林石，O.编），第405 - 451页，学术出版社，纽约）。
