title: Methylmalonate impairs mitochondrial respiration supported by NADH-linked substrates: involvement of mitochondrial glutamate metabolism.
abstract: The neurodegeneration that occurs in methylmalonic acidemia is proposed to be associated with impairment of mitochondrial oxidative metabolism resulting from methylmalonate (MMA) accumulation. The present study evaluated the effects of MMA on oxygen consumption by isolated rat brain mitochondria in the presence of NADH-linked substrates (α-ketoglutarate, citrate, isocitrate, glutamate, malate, and pyruvate). Respiration supported either by glutamate or glutamate plus malate was significantly inhibited by MMA (1-10 mM), whereas no inhibition was observed when a cocktail of NADH-linked substrates was used. Measurements of glutamate transport revealed that the inhibitory effect of MMA on respiration maintained by this substrate is not due to inhibition of its mitochondrial uptake. In light of this result, the effect of MMA on the activity of relevant enzymes involved in mitochondrial glutamate metabolism was investigated. MMA had minor inhibitory effects on glutamate dehydrogenase and aspartate aminotransferase, whereas α-ketoglutarate dehydrogenase was significantly inhibited by this metabolite (K(i) = 3.65 mM). Moreover, measurements of α-ketoglutarate transport and mitochondrial MMA accumulation indicated that MMA/α-ketoglutarate exchange depletes mitochondria from this substrate, which may further contribute to the inhibition of glutamate-sustained respiration. To study the effect of chronic in vivo MMA treatment on mitochondrial function, young rats were intraperitoneally injected with MMA. No significant difference was observed in respiration between isolated brain mitochondria from control and MMA-treated rats, indicating that in vivo MMA treatment did not lead to permanent mitochondrial respiratory defects. Taken together, these findings indicate that the inhibitory effect of MMA on mitochondrial oxidative metabolism can be ascribed to concurrent inhibition of specific enzymes and lower availability of respiratory substrates.
authors.affiliation: Departamento de Patologia Clínica, Faculdade de Ciências Médicas, Universidade Estadual de Campinas, Campinas, São Paulo, Brazil.
authors.forename: Daniela R
authors.initials: DR
authors.lastname: Melo
authors.forename: Sandra R
authors.initials: SR
authors.lastname: Mirandola
authors.forename: Nilson A
authors.initials: NA
authors.lastname: Assunção
authors.forename: Roger F
authors.initials: RF
authors.lastname: Castilho
mesh_terms: D027322:Amino Acid Transport System X-AG / Q000378:metabolism; D000704:Analysis of Variance; D000818:Animals; D000831:Animals, Newborn; D002262:Carboxy-Lyases / Q000378:metabolism; D002950:Citrate (si)-Synthase / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005969:Glutamate Dehydrogenase / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism*; D007655:Ketoglutarate Dehydrogenase Complex / Q000378:metabolism; D007656:Ketoglutaric Acids / Q000378:metabolism; D008764:Methylmalonic Acid / Q000378:metabolism / Q000494:pharmacology*; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000378:metabolism*; D010101:Oxygen Consumption / Q000187:drug effects; D016548:Prosencephalon / Q000187:drug effects / Q000648:ultrastructure; D051381:Rats; D017208:Rats, Wistar
chemical_list: D027322:Amino Acid Transport System X-AG; D007656:Ketoglutaric Acids; D009097:Multienzyme Complexes; D018698:Glutamic Acid; D008764:Methylmalonic Acid; D007655:Ketoglutarate Dehydrogenase Complex; D005969:Glutamate Dehydrogenase; D002950:Citrate (si)-Synthase; D002262:Carboxy-Lyases; C019747:aspartate 4-decarboxylase