Prado L, Fernández E, Weissbach U, Blanco G, Quirós L M, Braña A F, Méndez C, Rohr J, Salas J A
Departamento de Biología Funcional e Instituto Universitario de Biotecnología de Asturias (IUBA-CSIC), Universidad de Oviedo, 33006 Oviedo, Spain.
Chem Biol. 1999 Jan;6(1):19-30. doi: 10.1016/s1074-5521(99)80017-9.
Mithramycin is a member of the clinically important aureolic acid group of antitumor drugs that interact with GC-rich regions of DNA nonintercalatively. These drugs contain a chromophore aglycon that is derived from condensation of ten acetate units (catalyzed by a type II polyketide synthase). The aglycones are glycosylated at two positions with different chain length deoxyoligosaccharides, which are essential for the antitumor activity. During the early stages of mithramycin biosynthesis, tetracyclic intermediates of the tetracycline-type occur, which must be converted at later stages into the tricyclic glycosylated molecule, presumably through oxidative breakage of the fourth ring.
Two intermediates in the mithramycin biosynthetic pathway, 4-demethyl-premithramycinone and premithramycin B, were identified in a mutant lacking the mithramycin glycosyltransferase and methyltransferase genes and in the same mutant complemented with the deleted genes, respectively. Premithramycin B contains five deoxysugars moieties (like mithramycin), but contains a tetracyclic aglycon moiety instead of a tricyclic aglycon. We hypothesized that transcription of mtmOIV (encoding an oxygenase) was impaired in this strain, preventing oxidative breakage of the fourth ring of premithramycin B. Inactivating mtmOIV generated a mithramycin nonproducing mutant that accumulated premithramycin B instead of mithramycin. In vitro assays demonstrated that MtmOIV converted premithramycin B into a tricyclic compound.
In the late stages of mithramycin biosynthesis by Strepyomyces argillaceus, a fully glycosylated tetracyclic tetracycline-like intermediate (premithramycin B) is converted into a tricyclic compound by the oxygenase MtmOIV. This oxygenase inserts an oxygen (Baeyer-Villiger oxidation) and opens the resulting lactone. The following decarboxylation and ketoreduction steps lead to mithramycin. Opening of the fourth ring represents one of the last steps in mithramycin biosynthesis.
光神霉素是临床上重要的奥瑞酸类抗肿瘤药物成员之一，它以非嵌入方式与富含鸟嘌呤 - 胞嘧啶（GC）的DNA区域相互作用。这些药物含有一个发色团苷元，它由十个乙酸酯单元缩合而成（由II型聚酮合酶催化）。苷元在两个位置被不同链长的脱氧寡糖糖基化，这对其抗肿瘤活性至关重要。在光神霉素生物合成的早期阶段，会出现四环素型的四环中间体，这些中间体在后期必须通过第四环的氧化断裂转化为三环糖基化分子。
在一个缺失光神霉素糖基转移酶和甲基转移酶基因的突变体中，以及在分别用缺失基因进行互补的同一突变体中，鉴定出了光神霉素生物合成途径中的两种中间体，即4 - 去甲基 - 前光神霉素酮和前光神霉素B。前光神霉素B含有五个脱氧糖部分（与光神霉素一样），但含有一个四环苷元部分而非三环苷元。我们推测该菌株中mtmOIV（编码一种加氧酶）的转录受到损害，阻止了前光神霉素B第四环的氧化断裂。使mtmOIV失活产生了一个不产生光神霉素的突变体，该突变体积累前光神霉素B而非光神霉素。体外试验证明，MtmOIV将前光神霉素B转化为一种三环化合物。
在泥质链霉菌合成光神霉素的后期阶段，一种完全糖基化的四环类四环素样中间体（前光神霉素B）通过加氧酶MtmOIV转化为三环化合物。这种加氧酶插入一个氧原子（拜耳 - 维利格氧化反应）并打开生成的内酯。随后的脱羧和酮还原步骤生成光神霉素。第四环的打开代表了光神霉素生物合成中的最后步骤之一。
