Structure-function analyses of the G729R 2-oxoadipate dehydrogenase genetic variant associated with L-lysine metabolism disorder

Xu Zhang, Natalia S. Nemeria, João Leandro, Sander Houten, Michael Lazarus, Gary Gerfen, Oliver Ozohanics, Attila Ambrus, Balint Nagy, Roman Brukh, Frank Jordan

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6 Scopus citations


2-Oxoadipate dehydrogenase (E1a, also known as DHTKD1, dehydrogenase E1 and transketolase domain-containing protein 1) is a thiamin diphosphate-dependent enzyme and part of the 2-oxoadipate dehydrogenase complex (OADHc) in L-lysine catabolism. Genetic findings have linked mutations in the DHTKD1 gene to several metabolic disorders. These include alpha-aminoadipic and alpha-ketoadipic aciduria (AMOXAD), a rare disorder of L-lysine, Lhydroxylysine, and L-tryptophan catabolism, characterized by clinical presentations such as developmental delay, mild-to-severe intellectual disability, ataxia, epilepsy, and behavioral disorders that cannot be currently managed by available treatments. A heterozygous missense mutation, c.2185G>A (p. G729R), in DHTKD1 has been identified in most AMOXAD cases. Here, we report that the G729R E1a variant when assembled into OADHc in vitro displays a 50-fold decrease in catalytic efficiency for NADH production and significantly reduced rate of glutaryl-CoA production by dihydrolipoamide succinyltransferase (E2o). However, the G729R E1a substitution did not affect any of the three side-reactions associated solely with G729R E1a, prompting us to determine the structure-function effects of this mutation. A multipronged systematic analysis of the reaction rates in the OADHc pathway, supplemented with results from chemical cross-linking and hydrogen-deuterium exchange MS, revealed that the c.2185G>A DHTKD1 mutation affects E1a–E2o assembly, leading to impaired channeling of OADHc intermediates. Cross-linking between the C-terminal region of both E1a and G729R E1a with the E2o lipoyl and core domains suggested that correct positioning of the C-terminal E1a region is essential for the intermediate channeling. These findings may inform the development of interventions to counter the effects of pathogenic DHTKD1 mutations.

Original languageEnglish
Article number012761
JournalJournal of Biological Chemistry
Issue number23
StatePublished - Jun 2020


  • 2-oxoadipate dehydrogenase
  • 2-oxoadipate dehydrogenase complex
  • Alpha-aminoadipic
  • Alpha-ketoadipic aciduria (AMOXAD)
  • DHTKD1
  • Dehydrogenase E1 and transketolase domain-containing protein 1
  • Gene; [c.2185G>A (p.G729R)] DHTKD1 mutation
  • HD exchange and chemical cross-linking mass spectrometry
  • Protein conformation
  • Substrate channeling


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