Li Hua, Wang Lijuan, He Jing
State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070, China.
Wei Sheng Wu Xue Bao. 2012 Oct 4;52(10):1210-8.
A putative type II polyketide synthase (PKS) gene cluster (sah) was found in the genome sequence of Streptomyces sahachiroi ATCC 33158 by bioinformatics. In order to discover its biological function, we cloned the sah cluster and made functional analyses by gene knockout and heterologous expression.
METHODS & RESULTS: Annotation of the predicted open reading frames (ORFs) by protein-protein blast revealed that the sah cluster is highly homologous with the spore-pigment biosynthetic cluster (whiE) in S. coelicolor A3 (2), except an additional O-methyltransferase gene (sahI). Three genes of the sah cluster, sahG, sahH and sahI, which are putatively involved in post-PKS modifications, were respectively in-frame deleted by double crossover. We observed the color of the mutant strains spores apparently changed. HPLC and LC-MS analyses demonstrated that heterologous expression of the sah-minimal PKS gene and the whiE-minimal PKS gene in S. lividans ZX1 produced the same hydrophilic red pigments.
The sah cluster is responsible for the spore-pigment biosynthesis of S. sahachiroi ATCC 33158. It catalyzes the formation of polyketides with the same core structures as that of the whiE cluster.
通过生物信息学方法在酒井链霉菌ATCC 33158的基因组序列中发现了一个假定的II型聚酮合酶（PKS）基因簇（sah）。为了探究其生物学功能，我们克隆了sah基因簇，并通过基因敲除和异源表达进行功能分析。
通过蛋白质-蛋白质比对注释预测的开放阅读框（ORF），发现sah基因簇与天蓝色链霉菌A3（2）中的孢子色素生物合成基因簇（whiE）高度同源，只是多了一个O-甲基转移酶基因（sahI）。通过双交换分别对sah基因簇中可能参与聚酮合酶后修饰的三个基因sahG、sahH和sahI进行了框内缺失。我们观察到突变菌株孢子的颜色明显改变。高效液相色谱（HPLC）和液相色谱-质谱联用（LC-MS）分析表明，在变铅青链霉菌ZX1中异源表达sah最小聚酮合酶基因和whiE最小聚酮合酶基因产生了相同的亲水性红色色素。
sah基因簇负责酒井链霉菌ATCC 33158的孢子色素生物合成。它催化形成与whiE基因簇具有相同核心结构的聚酮化合物。
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