The Sinorhizobium meliloti MucR protein, which is essential for the production of high-molecular-weight succinoglycan exopolysaccharide, binds to short DNA regions upstream of exoH and exoY

Bertram-Drogatz PA, Quester I, Becker A, Pühler A. The Sinorhizobium meliloti MucR protein, which is essential for the production of high-molecular-weight succinoglycan exopolysaccharide, binds to short DNA regions upstream of exoH and exoY. MOLECULAR AND GENERAL GENETICS. 1998;257(4):433-441.Sinorhizobium meliloti (Rhizobium meliloti) is able to produce two different exopolysaccharides, succinoglycan and galactoglucan. Mutations in the mucR gene of S. meliloti result in the stimulation of galactoglucan synthesis, while the type of succinoglycan produced is modified. In culture supernatants of a mucR mutant, low-molecular-weight succinoglycan is present, whereas no high-molecular-weight succinoglycan could be detected. The biosynthesis of succinoglycan is directed by the products of the exo gene cluster. Two DNA fragments from this cluster, one located in front of the exoH gene and one in the intergenic region between the divergently transcribed genes exoX and exoY, were shown to represent effective binding sites for MucR. Whereas the latter binding site contains an inverted repeat motif, the former does not. However, the binding of MucR did not strongly modify the transcription of the exo genes involved. In the mucR mutant the expression levels of exoH-lacZ and exoX-lacZ transcriptional fusions were found to be increased 1.5- and 1.7-fold, respectively. On the other hand, the expression level of an exoY-lacZ transcriptional fusion was found to be 1.5-fold lower in the mucR mutant than in the wild-type background. Comparison of the DNA sequences of MucR-binding sites provides insight into the structural requirements for binding of MucR

The regulatory protein MucR binds to a short DNA region located upstream of the mucR coding region in Rhizobium meliloti

Bertram-Drogatz PA, Rüberg S, Becker A, Pühler A. The regulatory protein MucR binds to a short DNA region located upstream of the mucR coding region in Rhizobium meliloti. Mol Gen Genet. 1997;254(5):529-538.The Rhizobium meliloti MucR protein is known to regulate the biosynthesis of the two exopolysaccharides, succinoglycan and galactoglucan. The mucR gene was successfully overexpressed in Escherichia coli BL21 cells by heat shock induction using a two-plasmid system. Cell extracts of the production strain contained about 20% of a polypeptide of 17 kDa apparent molecular mass, corresponding to the size expected for MucR. As shown by an electrophoretic mobility shift assay, these extracts were active in the specific retardation of a 219-bp DNA fragment including 134-bp of the non-coding region upstream of the mucR gene. Primer extension analysis showed that this DNA fragment was located within the transcribed region upstream of the mucR gene. Competition experiments revealed that a 44-bp sequence present within the 134-bp upstream of the mucR gene contained the MucR binding site. Although binding of MucR to this site exhibited a moderate dissociation constant of K-d approximate to 1.4 x 10(-7) M, the reaction was highly specific since fragments containing binding sites for the homologous Ros protein from Agrobacterium tumefaciens were not able to compete for MucR binding

Regulation of succinoglycan and galactoglucan biosynthesis in Sinorhizobium meliloti

Becker A, Rüberg S, Baumgarth B, Bertram-Drogatz PA, Quester I, Pühler A. Regulation of succinoglycan and galactoglucan biosynthesis in Sinorhizobium meliloti. JOURNAL OF MOLECULAR MICROBIOLOGY AND BIOTECHNOLOGY. 2002;4(3):187-190.Sinorhizobium meliloti (Rhizobium meliloti) 2011 has the ability to produce the two acidic exopolysaccharides succinoglycan (EPSI) and galactoglucan (EPSII). EPSI is a branched heteropolysaccharide composed of octasaccharide repeating units, whereas EPSII is a linear heteropolysaccharide consisting of disaccharide subunits. The exo-exs and exp gene clusters are involved in the biosynthesis of EPSI and EPSII, respectively. EPSI and EPSII biosynthesis genes are differentially expressed resulting in a complex regulation of EPS production in S. meliloti. The phosphate concentration was identified as an Important factor affecting the expression of exp genes