Isomerization of -3-methylglutaconic acid.

3-Methylglutaconic (3MGC) aciduria is a common phenotypic feature of a growing number of inborn errors of metabolism. "Primary" 3MGC aciduria is caused by deficiencies in leucine pathway enzymes while "secondary" 3MGC aciduria results from inborn errors of metabolism that impact mitochondrial energy production. The metabolic precursor of 3MGC acid is -3MGC CoA, an intermediate in the leucine catabolism pathway. Gas chromatography-mass spectrometry (GC-MS) analysis of commercially available -3MGC acid yielded a mixture of and isomers while H-NMR spectroscopy of -3MGC acid at 25°C provided no evidence for the isomer. When -3MGC acid was incubated under conditions used for sample derivatization prior to GC-MS (but with no trimethylsilane added), H-NMR spectroscopy provided evidence of to isomerization. Incubation of -3MGC acid at 37°C resulted in time-dependent isomerization to -3MGC acid. -3MGC acid behaved in a similar manner except that, under identical incubation conditions, less isomerization occurred. In agreement with these experimental results, molecular modeling studies provided evidence that the energy minimized structure of -3MGC acid is 4 kJ/mol more stable than that for -3MGC acid. Once generated in vivo, -3MGC acid is proposed to isomerize via a mechanism involving π electron delocalization with formation of a resonance structure that permits bond rotation. The data presented are consistent with the occurrence of both diastereomers in urine samples of subjects with 3MGC aciduria.