Sinorhizobium melilotiMutants Lacking Phosphotransferase System Enzyme HPr or EIIA Are Altered in Diverse Processes, Including Carbon Metabolism, Cobalt Requirements, and Succinoglycan Production

Abstract

Sinorhizobium meliloti is a member of the Alphaproteobacteria that fixes nitrogen when it is in a symbiotic relationship. Genes for an incomplete phosphotransferase system (PTS) have been found in the genome of S. meliloti. The genes present code for Hpr and ManX (an EIIA Man -type enzyme). HPr and EIIA regulate carbon utilization in other bacteria. hpr and manX in-frame deletion mutants exhibited altered carbon metabolism and other phenotypes. Loss of HPr resulted in partial relief of succinate-mediated catabolite repression, extreme sensitivity to cobalt limitation, rapid die-off during stationary phase, and altered succinoglycan production. Loss of ManX decreased expression of melA-agp and lac, the operons needed for utilization of ␣-and ␤-galactosides, slowed growth on diverse carbon sources, and enhanced accumulation of high-molecularweight succinoglycan. A strain with both hpr and manX deletions exhibited phenotypes similar to those of the strain with a single hpr deletion. Despite these strong phenotypes, deletion mutants exhibited wild-type nodulation and nitrogen fixation when they were inoculated onto Medicago sativa. The results show that HPr and ManX (EIIA Man ) are involved in more than carbon regulation in S. meliloti and suggest that the phenotypes observed occur due to activity of HPr or one of its phosphorylated forms.

HPrK Regulates Succinate-Mediated Catabolite Repression in the Gram-Negative SymbiontSinorhizobium meliloti

“…Transport-related PTS proteins EIIB and EIIC are absent, indicating that PTS proteins that have been retained in S. meliloti probably provide critical regulatory functions even though they likely do not catalyze transport (4,29). In support of this hypothesis, recent work in S. meliloti has shown that the HPr and ManX proteins are involved in regulation of many aspects of cellular physiology, including carbon metabolism, catabolite repression, exopolysaccharide synthesis, and survival in stationary phase (44).…”

“…Growth curves and growth rates were obtained as explained previously (44). Concentration of single carbon sources was 0.4%, while concentrations for dual carbon sources were 0.05% for succinate and 0.1% for the secondary carbon source.…”

“…Experiments with a fraction of suppressors higher than 10% were not included in the results. Growth experiments for evaluation of gene expression were conducted with strains transformed with plasmid pCAP11, and fluorescence was measured as described previously (44).…”

An ABC-Type Cobalt Transport System Is Essential for Growth of Sinorhizobium melilotiat Trace Metal Concentrations

“…The S. meliloti BluB protein was recently shown to catalyze a missing step in vitamin B 12 synthesis by cannibalizing flavin to form 5,6-dimethylbenzimidazole, the lower ligand of vitamin B 12 (7,67). Recently, Co 2ϩ was observed to be required for survival of an S. meliloti mutant that lacks the phosphotransferase system enzyme Hpr (48). BioM (CbiO homolog) and BioN (CbiQ homolog) in S. meliloti have been reported to import biotin (18), and these along with BioY appear to represent an ECFtype biotin transporter.…”

RpoE2 of Sinorhizobium meliloti is necessary for trehalose synthesis and growth in hyperosmotic media

“…Significant differences in glycogen biosynthesis have been shown as a function of the metabolic state of the cells and the carbon source (Encarnació n et al, 2002;Tavernier et al, 1998). Precursors not only affect metabolic flux (Carpinelli et al, 2006) but also are input signals in regulatory circuits (Pinedo et al, 2008). Glycogen metabolism is connected to other major cellular processes by a complex regulatory network (Eydallin et al, 2007).…”