Schwab M, Reisenzein H, Högenauer G
Institut für Mikrobiologie, Universität Graz, Austria.
Mol Microbiol. 1993 Mar;7(5):795-803. doi: 10.1111/j.1365-2958.1993.tb01170.x.
Regulation of the traM gene, which encodes a factor essential for conjugation of resistance plasmid R1, was studied in vivo using translational gene fusion. traM"lacZ fusion constructs were transferred to the chromosome via the recombinant phage lambda RZ5. The level of beta-galactosidase expressed by the lysogens indicates that the traM promoters are very active. Expression of traM was diminished five- to sixfold when the single-copy plasmids R1 or R1-19 were present in trans. When recombinant plasmids carrying traM were present at higher copy numbers, traM expression was reduced as much as 45-fold. The negative effect of R1 plasmids on traM expression in trans, which we interpret as autoregulation, was observed regardless of whether the plasmids were conjugatively repressed or derepressed. Site-specific mutagenesis of the region encoding the N-terminus of the TraM protein eliminated the autoregulative effect indicating that the N-terminal amino acids of the protein are important to its DNA-binding function. The autoregulatory behaviour of TraM is allele specific. R1- or P307-encoded TraM molecules were found to recognize only the cognate DNA.
利用翻译基因融合技术在体内研究了编码耐药质粒R1接合所必需因子的traM基因的调控。traM"lacZ融合构建体通过重组噬菌体λRZ5转移到染色体上。溶原菌表达的β-半乳糖苷酶水平表明traM启动子非常活跃。当单拷贝质粒R1或R1-19反式存在时,traM的表达降低了五到六倍。当携带traM的重组质粒以更高拷贝数存在时,traM表达降低多达45倍。无论质粒是接合抑制还是去抑制,都观察到R1质粒对反式traM表达的负面影响,我们将其解释为自动调节。对TraM蛋白N端编码区域进行位点特异性诱变消除了自动调节作用,表明该蛋白的N端氨基酸对其DNA结合功能很重要。TraM的自动调节行为具有等位基因特异性。发现R1或P307编码的TraM分子仅识别同源DNA。