Disruption of a gene encoding a putative γ-butyrolactone-binding protein in Streptomyces tendae affects nikkomycin production
Disruption of a gene encoding a putative γ-butyrolactone-binding protein in Streptomyces tendae affects nikkomycin production
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DOI：
10.1007/s002530100621
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发表时间：
2001-08-01
影响因子：
5
通讯作者：
Cary, JW
中科院分区：
文献类型：
--
作者：
Engel, P;Scharfenstein, LL;Cary, JW
克隆并测序了来自野生型、产尼可霉素的链霉菌 ATCC 31160 菌株基因组的 2.6-kb BamHI 片段。这个 2.6-kb BamHI 片段对应于转座子 Tn4560 插入的 DNA 位点，以产生不产生尼可霉素的突变体。在这个 2.6 kb BamHI 片段中发现了一个可能的 660 个核苷酸的 ORF，其中每个密码子的第三个碱基在 92% 的密码子中是 G 或 C。由该 ORF（TarA，tendae 自动调节受体）编码的推导氨基酸序列与几种 γ-丁内酯结合蛋白显示出很强的同源性，这些结合蛋白负向调节其他链霉菌中的抗生素产生，并具有螺旋-转角-螺旋 DNA 结合基序。 tarA 编码螺旋-转角-螺旋基序的部分（179 个核苷酸）被 ermE 取代，野生型 S. tentae 被 pDH5（一种基因破坏载体）中携带的构建体转化。 Southern 杂交表明，ermE 已插入一个红霉素抗性分离株的 2.6-kb BamHI 区域。 Northern 杂交表明 tarA 破坏显着增加了破坏的 tarA mRNA 的量。这表明 TarA 负向调节其自身的合成。 tarA 破坏体产生的尼可霉素被延迟，但在生产培养基中孵育较长时间后达到野生型水平。
A 2.6-kb BamHI fragment from the genome of the wild-type, nikkomycin-producing strain of Streptomyces tendae ATCC 31160 was cloned and sequenced. This 2.6-kb BamHI fragment corresponds to the DNA site where transposon Tn4560 had inserted to create a nikkomycin-nonproducing mutant. A possible ORF of 660 nucleotides was found in this 2.6-kb BamHI fragment, in which the third base of each codon was either G or C in 92% of the codons. The deduced amino acid sequence coded by this ORF (TarA, tendae autoregulator receptor) shows strong homology with several gamma -butyrolactone-binding proteins that negatively regulate antibiotic production in other streptomycetes and have a helix-turn-helix DNA-binding motif. A portion (179 nucleotides) of tarA that encodes the helix-turn-helix motif was replaced with ermE, and wild-type S. tendae was transformed with this construct borne in pDH5, a gene-disruption vector. Southern hybridization indicated that ermE had inserted in the 2.6-kb BamHI region in one isolate that is erythromycin resistant. Northern hybridization indicated that tarA disruption significantly increased the amount of disrupted-tarA mRNA. This suggests that TarA negatively regulates its own synthesis. Nikkomycin production by the tarA disruptant was delayed but reached the wild-type level after longer incubation in production medium.