Funa N, Ohnishi Y, Fujii I, Shibuya M, Ebizuka Y, Horinouchi S
Department of Biotechnology, Graduate School of Agriculture and Life Sciences, University of Tokyo, Japan.
Nature. 1999 Aug 26;400(6747):897-9. doi: 10.1038/23748.
Chalcone synthases, which biosynthesize chalcones (the starting materials for many flavonoids), have been believed to be specific to plants. However, the rppA gene from the Gram-positive, soil-living filamentous bacterium Streptomyces griseus encodes a 372-amino-acid protein that shows significant similarity to chalcone synthases. Several rppA-like genes are known, but their functions and catalytic properties have not been described. Here we show that a homodimer of RppA catalyses polyketide synthesis: it selects malonyl-coenzyme-A as the starter, carries out four successive extensions and releases the resulting pentaketide to cyclize to 1,3,6,8-tetrahydroxynaphthalene (THN). Site-directed mutagenesis revealed that, as in other chalcone synthases, a cysteine residue is essential for enzyme activity. Disruption of the chromosomal rppA gene in S. griseus abolished melanin production in hyphae, resulting in 'albino' mycelium. THN was readily oxidized to form 2,5,7-trihydroxy-1,4-naphthoquinone (flaviolin), which then randomly polymerized to form various coloured compounds. THN formed by RppA appears to be an intermediate in the biosynthetic pathways for not only melanins but also various secondary metabolites containing a naphthoquinone ring. Therefore, RppA is a chalcone-synthase-related synthase that synthesizes polyketides and is found in the Streptomyces and other bacteria.
查尔酮合酶可生物合成查尔酮(许多类黄酮的起始原料),一直以来被认为是植物特有的。然而,革兰氏阳性、土壤生存的丝状细菌灰色链霉菌中的rppA基因编码一种372个氨基酸的蛋白质,该蛋白质与查尔酮合酶具有显著的相似性。已知有几个类似rppA的基因,但它们的功能和催化特性尚未被描述。在此,我们表明RppA的同型二聚体催化聚酮化合物的合成:它选择丙二酰辅酶A作为起始物,进行四次连续延伸,并释放出由此产生的五酮化合物以环化生成1,3,6,8 - 四羟基萘(THN)。定点诱变显示,与其他查尔酮合酶一样,一个半胱氨酸残基对酶活性至关重要。破坏灰色链霉菌染色体上的rppA基因会消除菌丝中的黑色素产生,导致形成“白化”菌丝体。THN很容易被氧化形成2,5,7 - 三羟基 - 1,4 - 萘醌(黄酮醇),然后随机聚合形成各种有色化合物。由RppA形成的THN似乎不仅是黑色素生物合成途径中的中间体,也是各种含有萘醌环的次生代谢产物生物合成途径中的中间体。因此,RppA是一种与查尔酮合酶相关的合酶,可合成聚酮化合物,存在于链霉菌和其他细菌中。