Molecular Studies for Putative Promoter Activity in mdh sucCDAB Operon in Sinorhizobium meliloti

Abstract

The theoretical data on the malate dehydrogenase (mdh), succinyl-CoA synthetase (sucCDAB) operon in Sinorhizobium meliloti and the experimental data suggest that this operon is regulated from the promoter upstream of mdh gene. On the other hand, the untranslated intergenic region between the sucD and sucA genes is TA rich and could also contains active promoter site, as it was described previously in Bradyrhizobium japonicum. In this study the mdh upstream region and the intergenic region between sucD and sucA were analyzed, isolated and cloned in pOT1-green fluorescence protein (gfp)-fusion plasmid to examine the possibility of promoter activity. Sequences analyzed by promoter hunter program indicated that both regions have promoter sequences and transcription starting sites (TSS). In addition the measurement of the relative fluorescence units (RFU) indicated that the mdh upstream region contains a constitutive promoter which was active under all tested conditions. While the intergenic region between sucD and sucA also contain active promoter site, induced only by LBmc medium and M9 medium containing glutamate as sole carbon source. The overall results suggest that sucA expression is initiated from its own upstream promoter.