Kempf B, Bremer E
Max-Planck-Institute for Terrestrial Microbiology, Marburg, Germany.
J Biol Chem. 1995 Jul 14;270(28):16701-13. doi: 10.1074/jbc.270.28.16701.
Exogenously provided glycine betaine can efficiently protect Bacillus subtilis from the detrimental effects of high osmolarity environments. Through functional complementation of an Escherichia coli mutant deficient in glycine betaine uptake with a gene library from B. subtilis, we have identified a multicomponent glycine betaine transport system, OpuA. Uptake of radiolabeled glycine betaine in B. subtilis was found to be osmotically stimulated and was strongly decreased in a mutant strain lacking the OpuA transport system. DNA sequence analysis revealed that the components of the OpuA system are encoded by anoperon (opuA) comprising three structural genes: opuAA, opuAB, and opuAC. The products of these genes exhibit features characteristic for binding protein-dependent transport systems and in particular show homology to the glycine betaine uptake system ProU from E. coli. Expression of the opuA operon is under osmotic control. The transcriptional initiation sites of opuA were mapped by high resolution primer extension analysis, and two opuA mRNAs were detected that differed by 38 base pairs at their 5' ends. Synthesis of the shorter transcript was strongly increased in cells grown at high osmolarity, whereas the amount of the longer transcript did not vary in response to medium osmolarity. Physical and genetic mapping experiments allowed the positioning the opuA operon at 25 degrees on the genetic map of B. subtilis.
外源提供的甘氨酸甜菜碱能够有效地保护枯草芽孢杆菌免受高渗环境的有害影响。通过用枯草芽孢杆菌的基因文库对缺乏甘氨酸甜菜碱摄取能力的大肠杆菌突变体进行功能互补，我们鉴定出了一个多组分甘氨酸甜菜碱转运系统OpuA。在枯草芽孢杆菌中发现，放射性标记的甘氨酸甜菜碱的摄取受到渗透压刺激，并且在缺乏OpuA转运系统的突变菌株中摄取量大幅下降。DNA序列分析表明，OpuA系统的组分由一个操纵子（opuA）编码，该操纵子包含三个结构基因：opuAA、opuAB和opuAC。这些基因的产物表现出依赖结合蛋白的转运系统的特征，尤其与大肠杆菌的甘氨酸甜菜碱摄取系统ProU具有同源性。opuA操纵子的表达受渗透压控制。通过高分辨率引物延伸分析确定了opuA的转录起始位点，检测到两种opuA mRNA，它们在5'端相差38个碱基对。较短转录本的合成在高渗条件下生长的细胞中显著增加，而较长转录本的量不随培养基渗透压变化。物理和遗传作图实验将opuA操纵子定位在枯草芽孢杆菌遗传图谱的25度处。
