Characterization of Mutations That Affect the Nonoxidative Pentose Phosphate Pathway in Sinorhizobium meliloti

Sinorhizobium meliloti is a Gram-negative alphaproteobacterium that can enter into a symbiotic relationship withMedicago sativa andMedicago truncatula . Previous work determined that a mutation in thetkt2 gene, which encodes a putative transketolase, could prevent medium acidification associated with a mutant strain unable to metabolize galactose. Since the pentose phosphate pathway inS. meliloti is not well studied, strains carrying mutations in eithertkt2 andtal , which encodes a putative transaldolase, were characterized. Carbon metabolism phenotypes revealed that both mutants were impaired in growth on erythritol and ribose. This phenotype was more pronounced for thetkt2 mutant strain, which also displayed auxotrophy for aromatic amino acids. Changes in pentose phosphate pathway metabolite concentrations were also consistent with a mutation in eithertkt2 ortal . The concentrations of metabolites in central carbon metabolism were also found to shift dramatically in strains carrying atkt2 mutation. While the concentrations of proteins involved in central carbon metabolism did not change significantly under any conditions, the levels of those associated with iron acquisition increased in the wild-type strain with erythritol induction. These proteins were not detected in either mutant, resulting in less observable rhizobactin production in thetkt2 mutant. While both mutants were impaired in succinoglycan synthesis, only thetkt2 mutant strain was unable to establish symbiosis with alfalfa. These results suggest thattkt2 andtal play central roles in regulating the carbon flow necessary for carbon metabolism and the establishment of symbiosis.IMPORTANCE Sinorhizobium meliloti is a model organism for the study of plant-microbe interactions and metabolism, especially because it effects nitrogen fixation. The ability to derive the energy necessary for nitrogen fixation is dependent on an organism's ability to metabolize carbon efficiently. The pentose phosphate pathway is central in the interconversion of hexoses and pentoses. This study characterizes the key enzymes of the nonoxidative branch of the pentose phosphate pathway by using defined genetic mutations and shows the effects the mutations have on the metabolite profile and on physiological processes such as the biosynthesis of exopolysaccharide, as well as the ability to regulate iron acquisition.