The E2 domain of OdhA of Corynebacterium glutamicum has succinyltransferase activity dependent on lipoyl residues of the acetyltransferase AceF.
Oxoglutarate dehydrogenase (ODH) and pyruvate dehydrogenase (PDH) complexes catalyze key reactions in central metabolism, and in Corynebacterium glutamicum there is indication of an unusual supercomplex consisting of AceE (E1), AceF (E2), and Lpd (E3) together with OdhA. OdhA is a fusion protein of additional E1 and E2 domains, and odhA orthologs are present in all Corynebacterineae, including, for instance, Mycobacterium tuberculosis. Here we show that deletion of any of the individual domains of OdhA in C. glutamicum resulted in loss of ODH activity, whereas PDH was still functional. On the other hand, deletion of AceF disabled both PDH activity and ODH activity as well, although isolated AceF protein had solely transacetylase activity and no transsuccinylase activity. Surprisingly, the isolated OdhA protein was inactive with 2-oxoglutarate as the substrate, but it gained transsuccinylase activity upon addition of dihydrolipoamide. Further enzymatic analysis of mutant proteins and mutant cells revealed that OdhA specifically catalyzes the E1 and E2 reaction to convert 2-oxoglutarate to succinyl-coenzyme A (CoA) but fully relies on the lipoyl residues provided by AceF involved in the reactions to convert pyruvate to acetyl-CoA. It therefore appears that in the putative supercomplex in C. glutamicum, in addition to dihydrolipoyl dehydrogenase E3, lipoyl domains are also shared, thus confirming the unique evolutionary position of bacteria such as C. glutamicum and M. tuberculosis.
Hoffelder
Melanie
M
Institute of Biotechnology, Forschungszentrum Jülich, Leo-Brandt-Str. 1, D-52425 Jülich, Germany.
Raasch
Katharina
K
van Ooyen
Jan
J
Eggeling
Lothar
L
D000123:Acetyltransferases / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D048230:Corynebacterium glutamicum / Q000201:enzymology* / Q000235:genetics; D057075:Enzyme Assays; D009154:Mutation; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism; D016133:Polymerase Chain Reaction
D001426:Bacterial Proteins; D010088:Oxidoreductases; D000123:Acetyltransferases