Off-pathway, oxygen-dependent thiamine radical in the Krebs cycle.

TitleOff-pathway, oxygen-dependent thiamine radical in the Krebs cycle.
Publication TypeJournal Article
Year of Publication2008
AuthorsFrank, RAW, Kay, CWM, Hirst, J, Luisi, BF
JournalJ Am Chem Soc
Date Published2008 Feb 06
KeywordsCatalysis, Citric Acid Cycle, Electron Spin Resonance Spectroscopy, Free Radicals, Geobacillus stearothermophilus, Models, Molecular, Molecular Structure, Oxygen, Peroxides, Thiamine

The catalytic cofactor thiamine diphosphate is found in many enzymes of central metabolism and is essential in all extant forms of life. We demonstrate the presence of an oxygen-dependent free radical in the thiamine diphosphate-dependent Escherichia coli 2-oxoglutarate dehydrogenase, which is a key component of the tricarboxylic acid (Krebs) cycle. The radical was sufficiently long-lived to be trapped by freezing in liquid nitrogen, and its electronic structure was investigated by electron paramagnetic resonance (EPR) and electron-nuclear double resonance (ENDOR). Taken together, the spectroscopic results revealed a delocalized pi radical on the enamine-thiazolium intermediate within the enzyme active site. The radical is generated as an intermediate during substrate turnover by a side reaction with molecular oxygen, resulting in the continuous production of reactive oxygen species under aerobic conditions. This off-pathway reaction may account for metabolic dysfunction associated with several neurodegenerative diseases. The possibility that the on-pathway reaction may proceed via a radical mechanism is discussed.

Alternate JournalJ. Am. Chem. Soc.
Citation Key10.1021/ja076468k
PubMed ID18183975
Grant ListMC_U105663141 / / Medical Research Council / United Kingdom
/ / Wellcome Trust / United Kingdom