Keller N P, Kantz N J, Adams T H
Department of Plant Pathology and Microbiology, Texas A&M University, College Station 77843.
Appl Environ Microbiol. 1994 May;60(5):1444-50. doi: 10.1128/aem.60.5.1444-1450.1994.
Aspergillus nidulans produces the carcinogenic mycotoxin sterigmatocystin (ST), the next-to-last precursor in the aflatoxin (AF) biosynthetic pathway found in the closely related fungi Aspergillus flavus and Aspergillus parasiticus. We identified and characterized an A. nidulans gene, verA, that is required for converting the AF precursor versicolorin A to ST. verA is closely related to several polyketide biosynthetic genes involved in polyketide production in Streptomyces spp. and exhibits extended sequence similarity to A. parasiticus ver-1, a gene proposed to encode an enzyme involved in converting versicolorin A to ST. By performing a sequence analysis of the region 3' to verA, we identified two additional open reading frames, designated ORF1 and ORF2. ORF2 is closely related to a number of cytochrome P-450 monooxygenases, while ORF1 shares identity with the gamma subunit of translation elongation factor 1. Given that several steps in the ST-AF pathway may require monooxygenase activity and that AF biosynthetic genes are clustered in A. flavus and A. parasiticus, we suggest that verA may be part of a cluster of genes required for ST biosynthesis. We disrupted the verA coding region by inserting the A. nidulans argB gene into the center of the coding region and transformed an A. nidulans argB2 mutant to arginine prototrophy. Seven transformants that produced DNA patterns indicative of a verA disruption event were grown under ST-inducing conditions, and all of the transformants produced versicolorin A but negligible amounts of ST (200-fold to almost 1,000-fold less than the wild type), confirming the hypothesis that verA encodes an enzyme necessary for converting versicolorin A to ST.
构巢曲霉产生致癌霉菌毒素柄曲霉素（ST），它是在密切相关的真菌黄曲霉和寄生曲霉中发现的黄曲霉毒素（AF）生物合成途径中的倒数第二个前体。我们鉴定并表征了构巢曲霉的一个基因verA，它是将AF前体杂色曲霉素A转化为ST所必需的。verA与链霉菌属中参与聚酮化合物产生的几个聚酮化合物生物合成基因密切相关，并且与寄生曲霉的ver-1具有延伸的序列相似性，ver-1是一个被认为编码参与将杂色曲霉素A转化为ST的酶的基因。通过对verA 3'端区域进行序列分析，我们鉴定出另外两个开放阅读框，分别命名为ORF1和ORF2。ORF2与许多细胞色素P-450单加氧酶密切相关，而ORF1与翻译延伸因子1的γ亚基具有同一性。鉴于ST-AF途径中的几个步骤可能需要单加氧酶活性，并且AF生物合成基因在黄曲霉和寄生曲霉中是成簇的，我们认为verA可能是ST生物合成所需的基因簇的一部分。我们通过将构巢曲霉的argB基因插入编码区中心来破坏verA编码区，并将构巢曲霉的argB2突变体转化为精氨酸原养型。在ST诱导条件下培养了七个产生指示verA破坏事件的DNA模式的转化体，所有转化体都产生了杂色曲霉素A，但产生的ST量可忽略不计（比野生型少200倍至近1000倍），这证实了verA编码将杂色曲霉素A转化为ST所必需的酶的假设。
