Fungal metabolic model for type I 3-methylglutaconic aciduria.

Aspergillus nidulans catabolizes Leu to acetyl-CoA and acetoacetate through a pathway homologous to that used by humans. Fungal hlyA encodes a bifunctional polypeptide comprising the last two enzymes in this pathway, 3-methylglutaconyl-CoA hydratase and 3-hydroxy-3-methylglutaryl-CoA lyase. hlyA transcription is specifically induced by Leu. A Delta hlyA mutation removing the complete 3-methylglutaconyl-CoA hydratase C-terminal domain prevents growth on Leu but not on lactose or other amino acids and, in agreement with the predicted enzyme function, leads to Leu-dependent accumulation of 3-methylglutaconic acid in the culture supernatant. These data represent a formal demonstration in vivo of the specific involvement of 3-methylglutaconyl-CoA hydratase in Leu catabolism. Type I 3-methylglutaconic aciduria patients deficient in 3-methylglutaconyl-CoA hydratase show urinary excretion of 3-methylglutaconic acid and, in contrast to the other three types of methylglutaconic acidurias, 3-hydroxyisovaleric acid excretion. Gas chromatography-mass spectrometry analysis revealed an accumulation of both diagnostic compounds in Delta hlyA culture supernatants, illustrating that the metabolic consequences of equivalent inborn errors of metabolism are conserved from fungi to humans. Using our fungal type I 3-methylglutaconic aciduria model, we show that metabolites accumulating in the deficient strain are toxic, although less so than those accumulating in a Delta mccB strain deficient for the upstream enzyme 3-methylcrotonyl-CoA carboxylase. Diagnostic metabolite accumulation is Leu concentration-dependent, in agreement with the ability of Leu intake restriction to reduce the levels of offending metabolites. Delta mccB and Delta hlyA mutations show additive Leu toxicities. The double mutant accumulates 3-methylglutaconic acid, which can therefore be synthesized through 3-methylcrotonyl-CoA carboxylase-dependent and -independent reactions.