Sinorhizobium meliloti dctA Mutants with Partial Ability To Transport Dicarboxylic Acids

AUTOR(ES)

Yurgel, Svetlana N.

FONTE

American Society for Microbiology

RESUMO

Sinorhizobium meliloti dctA encodes a transport protein needed for a successful nitrogen-fixing symbiosis between the bacteria and alfalfa. Using the toxicity of the DctA substrate fluoroorotic acid as a selective agent in an iterated selection procedure, four independent S. meliloti dctA mutants were isolated that retained some ability to transport dicarboxylates. Two mutations were located in a region called motif B located in a predicted transmembrane helix of the protein that has been shown in other members of the glutamate transporter family to be involved in cation binding. A G114D mutation was located in the third transmembrane helix, which had not previously been directly implicated in transport. Multiple sequence alignment of more than 60 members of the glutamate transporter family revealed a glycine at this position in nearly all members of the family. The fourth mutant was able to transport succinate at almost wild-type levels but was impaired in malate and fumarate transport. It contains two mutations: one in a periplasmic domain and the other predicted to be in the cytoplasm. Separation of the mutations showed that each contributed to the altered substrate preference. dctA deletion mutants that contain the mutant dctA alleles on a plasmid can proceed further in symbiotic development than null mutants of dctA, but none of the plasmids could support symbiotic nitrogen fixation, although they can transport dicarboxylates, some at relatively high levels.

Documentos Relacionados

• Roles of DctA and DctB in Signal Detection by the Dicarboxylic Acid Transport System of Rhizobium leguminosarum
• Dicarboxylic acid transport in Bradyrhizobium japonicum: use of Rhizobium meliloti dct gene(s) to enhance nitrogen fixation.
• Rhizobium meliloti and Rhizobium leguminosarum dctD gene products bind to tandem sites in an activation sequence located upstream of sigma 54-dependent dctA promoters.
• Identification and sequence analysis of the Rhizobium meliloti dctA gene encoding the C4-dicarboxylate carrier.
• Negative regulation of sigma 54-dependent dctA expression by the transcriptional activator DctD.