THIOSTREPTON-INDUCED GENE-EXPRESSION IN STREPTOMYCES-LIVIDANS
THIOSTREPTON-INDUCED GENE-EXPRESSION IN STREPTOMYCES-LIVIDANS
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DOI：
10.1128/jb.171.3.1459-1466.1989
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发表时间：
1989-03-01
影响因子：
3.2
通讯作者：
THOMPSON, CJ
中科院分区：
文献类型：
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作者：
MURAKAMI, T;HOLT, TG;THOMPSON, CJ
硫链丝菌素诱导变铅青链霉菌中四种功能未知的蛋白质（17，19，30和56千道尔顿）的表达。对编码19千道尔顿蛋白（tipA）的染色体基因进行了克隆和测序。tipA启动子的转录被硫链丝菌素诱导至少200倍。tipA转录起始位点（通过S1作图和引物延伸实验定位）之前是45个碱基对的不完全反向重复序列，其包括启动子的-10和-35区域。在体内非诱导条件下，这可能形成不被RNA聚合酶识别的十字形结构。将包含该区域的143个碱基对片段克隆到启动子探针载体pIJ486中。在该质粒pAK114中，硫链丝菌素诱导型tipA启动子控制编码氨基糖苷磷酸转移酶的卡那霉素抗性基因的表达。只有1 ng的硫链丝菌素被发现在S. pAK114诱导卡那霉素抗性生长。其他硫链菌素类抗生素也诱导tipA，但结构无关的抗生素抑制翻译没有影响。in s. lividans中，该启动子在生长期或稳定期均可被硫链丝菌素诱导。tipA启动子应该是在链霉菌中表达研究的有价值的工具。
Thiostrepton induced the expression of four proteins (17, 19, 30, and 56 kilodaltons) of unknown function in Streptomyces lividans. The chromosomal gene which encoded the 19-kilodalton protein (tipA) was cloned and sequenced. Transcription of the tipA promoter was induced at least 200-fold by thiostrepton. The tipA transcriptional start site (located by S1 mapping and primer extension experiments) was preceded by a 45-base-pair imperfect inverted-repeat sequence which included the -10 and -35 regions of the promoter. Under noninducing conditions in vivo, this might form a cruciform structure which is not recognize by RNA polymerase. A 143-base-pair fragment including this region was cloned into a promoter probe vector, pIJ486. In this plasmid, pAK114, the thiostrepton-inducible tipA promoter controlled the expression of a kanamycin resistance gene encoding an aminoglycoside phosphotransferase. As little as 1 ng of thiostrepton spotted on a lawn of S. lividans(pAK114) induced kanamycin-resistant growth. Other thiostreptonlike antibiotics also induced tipA, but structurally unrelated antibiotics which inhibit translation had no effect. In S. lividans, the promoter could be induced by thiostrepton during either growth or stationary phase. The tipA promoter should be a valuable tool for expression studies in streptomycetes.