Palaniappan Nadaraj, Alhamadsheh Mamoun M, Reynolds Kevin A
Department of Chemistry, Portland State University, Portland, OR 97297, USA.
J Am Chem Soc. 2008 Sep 17;130(37):12236-7. doi: 10.1021/ja8044162. Epub 2008 Aug 21.
The antifungal phoslactomycins (PLM A-F), produced by Streptomyces sp. HK803, are structurally unusual in that three of their four double bonds are in the cis form (Delta12,13, Delta14,15, Delta2,3). The PLM polyketide synthase (PKS) has the predicted dehydratase catalytic domain in modules 1, 2, and 5 required for establishing two of these cis double bonds (Delta12,13, Delta14,15), as well as the only trans Delta6,7 double bond. By contrast, the formation of the cis Delta2,3 in the unsaturated lactone moiety of PLMs has presented an enigma because the predicted dehydratase domain in module 7 is absent. Herein, we have demonstrated that the plmT2 gene product, with no homology to PKS dehydratase domains, is required for efficient formation of the cis Delta2,3 alkene. A series of new PLM products in which the C3 hydroxyl group is retained are made in plmT2 deletion mutants. In all of these cases, however, the hydroxyl group is esterified with malonic acid. These malonylated PLM products are converted to the corresponding cis Delta2,3 PLM products and acetic acid by a facile base-catalyzed decarboxylative elimination reaction. Complete or partial restoration of natural PLM production in a plmT2 deletion mutant can be accomplished by plasmid based expression of plmT2 or fos ORF4 (a homologous gene from the fostriecin biosynthetic gene cluster), respectively. The data indicate that dehydratase-independent pathways also function in establishment of unsaturated 6-membered lactone moieties in other PKS pathways and provide the first biosynthetic insights into the possible routes by which unusual malonylated polyketide products are generated.
由链霉菌属HK803产生的抗真菌磷乳霉素（PLM A - F）在结构上不同寻常，因为其四个双键中的三个呈顺式构型（Δ12,13、Δ14,15、Δ2,3）。PLM聚酮合酶（PKS）在模块1、2和5中具有预测的脱水酶催化结构域，这些结构域是形成其中两个顺式双键（Δ12,13、Δ14,15）以及唯一的反式Δ6,7双键所必需的。相比之下，PLMs不饱和内酯部分中顺式Δ2,3的形成一直是个谜，因为模块7中预测的脱水酶结构域缺失。在此，我们证明了plmT2基因产物与PKS脱水酶结构域无同源性，但它是顺式Δ2,3烯烃高效形成所必需的。在plmT2缺失突变体中产生了一系列保留C3羟基的新PLM产物。然而，在所有这些情况下，羟基都被丙二酸酯化。这些丙二酰化的PLM产物通过简便的碱催化脱羧消除反应转化为相应的顺式Δ2,3 PLM产物和乙酸。通过基于质粒表达plmT2或fos ORF4（来自磷霉素生物合成基因簇的同源基因），分别可以使plmT2缺失突变体中天然PLM的产生完全或部分恢复。数据表明，不依赖脱水酶的途径在其他PKS途径中不饱和六元内酯部分的形成中也起作用，并首次对异常丙二酰化聚酮产物产生的可能途径提供了生物合成方面的见解。
