Cloning and in vivo functional analysis by disruption of a gene encoding the γ-butyrolactone autoregulator receptor from Streptomyces natalensis
Cloning and in vivo functional analysis by disruption of a gene encoding the γ-butyrolactone autoregulator receptor from Streptomyces natalensis
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DOI：
10.1007/s00203-005-0047-7
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发表时间：
2005-12-01
影响因子：
2.8
通讯作者：
Hwang, YI
中科院分区：
文献类型：
--
作者：
Lee, KM;Lee, CK;Hwang, YI
编码γ -丁内酯自调节受体的基因，具有dna结合转录抑制因子的共同活性，控制链霉菌的次生代谢和/或形态分化，从纳他霉素生产链霉菌中克隆。链霉菌自调节受体BarA、FarA、ScbR和ArpA这两个高度保守区域的引物，PCR结果为102 bp。该条带序列与受体基因的预期区域高度相似。以102 bp插入物为探针，通过基因组Southern杂交，获得了一个687 bp的完整受体基因（sngR）。为了明确sngR在体内的功能，构建了sngR破坏菌株，并将其表型与野生型菌株进行了比较。sngr干扰菌比野生型菌株早6小时开始生产纳他霉素，其纳他霉素产量高出4.6倍。与野生菌株相比，该菌株孢子形成时间早，孢子数量多10倍。通过与完整的sngR互补，所有表型都恢复到野生型菌株的原始表型，表明s.a natalensis的自调节受体蛋白在纳他霉素的生物合成和产孢过程中都起着主要的负调节作用。
A gene encoding a gamma-butyrolactone autoregulator receptor, which has a common activity as DNA-binding transcriptional repressors controlling secondary metabolism and/or morphological differentiation in Streptomyces, was cloned from a natamycin producer, Streptomyces natalensis. PCR using the primers designed for the two highly conserved regions of Streptomyces autoregulator receptors (BarA, FarA, ScbR, and ArpA) gave a 102-bp band. The sequence of this band had a high similarity to the expected region of a receptor gene. By genomic Southern hybridization with the 102-bp insert as a probe, a 687-bp intact receptor gene (sngR) was obtained from S. natalensis. To clarify the in vivo function of sngR, a sngR-disrupted strain was constructed, and the phenotypes were compared with those of the wild-type strain. The sngR-disruptants started natamycin production 6 h earlier and showed a 4.6-fold higher production of natamycin than the wild-type strain. In addition, the sporulation began earlier and the number of spores was tenfold more abundant than that of the wild-type strain. All the phenotypes were restored back to the original phenotypes of the wild-type strain by complementation with the intact sngR, indicating that the autoregulator receptor protein of S. natalensis acts as a primary negative regulator both on the biosynthesis of natamycin and sporulation.