Catabolism of alpha-ketoglutarate by a sucA mutant of Bradyrhizobium japonicum: evidence for an alternative tricarboxylic acid cycle.
A complete tricarboxylic acid (TCA) cycle is generally considered necessary for energy production from the dicarboxylic acid substrates malate, succinate, and fumarate. However, a Bradyrhizobium japonicum sucA mutant that is missing alpha-ketoglutarate dehydrogenase is able to grow on malate as its sole source of carbon. This mutant also fixes nitrogen in symbiosis with soybean, where dicarboxylic acids are its principal carbon substrate. Using a flow chamber system to make direct measurements of oxygen consumption and ammonium excretion, we confirmed that bacteroids formed by the sucA mutant displayed wild-type rates of respiration and nitrogen fixation. Despite the absence of alpha-ketoglutarate dehydrogenase activity, whole cells of the mutant were able to decarboxylate alpha-[U-(14)C]ketoglutarate and [U-(14)C]glutamate at rates similar to those of wild-type B. japonicum, indicating that there was an alternative route for alpha-ketoglutarate catabolism. Because cell extracts from B. japonicum decarboxylated [U-(14)C]glutamate very slowly, the gamma-aminobutyrate shunt is unlikely to be the pathway responsible for alpha-ketoglutarate catabolism in the mutant. In contrast, cell extracts from both the wild type and mutant showed a coenzyme A (CoA)-independent alpha-ketoglutarate decarboxylation activity. This activity was independent of pyridine nucleotides and was stimulated by thiamine PP(i). Thin-layer chromatography showed that the product of alpha-ketoglutarate decarboxylation was succinic semialdehyde. The CoA-independent alpha-ketoglutarate decarboxylase, along with succinate semialdehyde dehydrogenase, may form an alternative pathway for alpha-ketoglutarate catabolism, and this pathway may enhance TCA cycle function during symbiotic nitrogen fixation.
Green
L S
LS
Biochemistry Department, University of Missouri, Columbia, Missouri 65211, USA. greenl@missouri.edu
Li
Y
Y
Emerich
D W
DW
Bergersen
F J
FJ
Day
D A
DA
D000445:Aldehyde Oxidoreductases / Q000378:metabolism*; D020369:Bradyrhizobium / Q000201:enzymology* / Q000235:genetics; D002262:Carboxy-Lyases / Q000378:metabolism*; D002952:Citric Acid Cycle; D003653:Decarboxylation; D017353:Gene Deletion; D005968:Glutamate Decarboxylase / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism; D007655:Ketoglutarate Dehydrogenase Complex / Q000235:genetics / Q000378:metabolism*; D007656:Ketoglutaric Acids / Q000378:metabolism*; D016296:Mutagenesis; D009586:Nitrogen Fixation; D050644:Succinate-Semialdehyde Dehydrogenase
D007656:Ketoglutaric Acids; D018698:Glutamic Acid; D000445:Aldehyde Oxidoreductases; D050644:Succinate-Semialdehyde Dehydrogenase; D007655:Ketoglutarate Dehydrogenase Complex; D002262:Carboxy-Lyases; D005968:Glutamate Decarboxylase; C021474:2-oxoglutarate decarboxylase