Transcriptional fusion mutants and Tn5-B20 transposon mutants were isolated where the only genes affected are believed to either be involved in the hypothetical GABA shunt or code for subunits of the alpha-ketoglutarate dehydrogenase enzyme complex of Sinorhizobium meliloti. The growth phenotypes of Rm30222 (gabT) and eight mutants in gabD1, 2, 3, and 4 on minimal media were comparable to that of the wild-type. Compared to wild-type, Rm30222 (gabT) lacked alpha-ketoglutarate-dependent gamma-aminobutyrate transaminase activity showed high induction of gabT on GABA but produced green plants indicative of being Fix+. Mutants in gabD alleles maintained wild-type levels of succinic semialdehyde dehydrogenase activity and could fix nitrogen as well as the wild-type in symbiosis. / Mutation of sucB encoding a subunit of a-ketoglutarate dehydrogenase produced a mutant, Rm30230, that initially had difficulty growing on minimal media supplemented with either arabinose or glutamate. In symbiosis with alfalfa, Rm30230 had a fix- phenotype and was also devoid of alpha-ketoglutarate dehydrogenase activity. The ability of Rm30230 to grow on arabinose or glutamate, without alpha-ketoglutarate dehydrogenase activity, strengthens the hypothesis that S. meliloti has a functional GABA shunt allowing it to circumvent the forward-direction TCA cycle from alpha-ketoglutarate to succinate. Mutation of the second potential dihydrolipoamide succinyltransferase component (E2) of alpha-ketoglutarate dehydrogenase yielded Rm30267 (SMb20019) with wild-type growth on minimal media and a Fix+ phenotype in plants. The introduction or a sucB mutation into the SMb20019 mutant background (Rm30275) was comparable to the sole sucB mutation. This finding shows that the locus SMb20019 cannot be substituted for sucB in the alpha-ketoglutarate dehydrogenase enzyme complex.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.111551 |
| Date | January 2008 |
| Creators | Trottier, Oliver. |
| Publisher | McGill University |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Format | application/pdf |
| Coverage | Master of Science (Department of Natural Resource Sciences.) |
| Rights | All items in eScholarship@McGill are protected by copyright with all rights reserved unless otherwise indicated. |
| Relation | alephsysno: 003134967, proquestno: AAIMR66731, Theses scanned by UMI/ProQuest. |
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