Dataset Information

Genetic analysis of the Sinorhizobium meliloti BacA protein: differential effects of mutations on phenotypes.

ABSTRACT: Sinorhizobium meliloti strains lacking BacA function are impaired in symbiosis with alfalfa host plants and display altered sensitivities to a number of compounds relative to wild-type strains. With the goal of finding clues to the currently unknown biological function(s) of BacA, we carried out a genetic analysis to determine which amino acids are critical for protein function and to attempt to ascertain whether the multiple phenotypes that result from a bacA-null allele were the result of a common cause or whether BacA has multiple functions. We have created a set of 20 site-directed mutants in which selected individual amino acids in bacA were replaced with glycine residues. The resulting mutants were characterized to determine how the various amino acid changes affected a number of phenotypes associated with loss of BacA function. Mutants H165G, W182G, D198G, and R284G had null phenotypes for all functions assayed, while mutants W57G, S83G, S231G, and K350G were indistinguishable from wild-type strains. The remaining 12 site-directed mutants demonstrate mixed phenotypic characteristics and fall into a number of distinctly different groups. These observations may be consistent with a role for BacA in multiple, nonoverlapping functions.

SUBMITTER: LeVier K

PROVIDER: S-EPMC100141 | biostudies-literature | 2001 Nov

REPOSITORIES: biostudies-literature

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Genetic analysis of the Sinorhizobium meliloti BacA protein: differential effects of mutations on phenotypes.

LeVier K K Walker G C GC

Journal of bacteriology 20011101 21

Sinorhizobium meliloti strains lacking BacA function are impaired in symbiosis with alfalfa host plants and display altered sensitivities to a number of compounds relative to wild-type strains. With the goal of finding clues to the currently unknown biological function(s) of BacA, we carried out a genetic analysis to determine which amino acids are critical for protein function and to attempt to ascertain whether the multiple phenotypes that result from a bacA-null allele were the result of a co ...[more]

PMID: 11591690

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Project description:Sinorhizobium meliloti is a Gram-negative alphaproteobacterium that can enter into a symbiotic relationship with Medicago sativa and Medicago truncatula Previous work determined that a mutation in the tkt2 gene, which encodes a putative transketolase, could prevent medium acidification associated with a mutant strain unable to metabolize galactose. Since the pentose phosphate pathway in S. meliloti is not well studied, strains carrying mutations in either tkt2 and tal, 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 the tkt2 mutant strain, which also displayed auxotrophy for aromatic amino acids. Changes in pentose phosphate pathway metabolite concentrations were also consistent with a mutation in either tkt2 or tal The concentrations of metabolites in central carbon metabolism were also found to shift dramatically in strains carrying a tkt2 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 the tkt2 mutant. While both mutants were impaired in succinoglycan synthesis, only the tkt2 mutant strain was unable to establish symbiosis with alfalfa. These results suggest that tkt2 and tal play central roles in regulating the carbon flow necessary for carbon metabolism and the establishment of symbiosis.IMPORTANCESinorhizobium 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.

Dataset Information

Genetic analysis of the Sinorhizobium meliloti BacA protein: differential effects of mutations on phenotypes.

Publications

Genetic analysis of the Sinorhizobium meliloti BacA protein: differential effects of mutations on phenotypes.

Similar Datasets

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