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Summary: l-Alloisoleucine formation froml-isoleucine was studiedin vitro andin vivo. When a healthy subject was loaded withl-isoleucine, plasma levels ofl-isoleucine and 3-methyl-2-oxopentanoate (KMV), as well asl-alloisoleucine, increased. Peak values were reached successively and were in the orderl-isoleucine > > KMV > >l-alloisoleucine. Metabolic clearance ofl-isoleucine and KMV was rapid; clearance ofl-alloisoleucine was considerably delayed. When human skin fibroblast cultures were challenged withl-isoleucine, KMV accumulated at a gradually decreased rate, whereasl-alloisoleucine accumulated at a gradually accelerated rate. KMV andl-alloisoleucine formation were related and depended on thel-isoleucine concentration applied. In cell lines derived from MSUD patients (classical form), metabolite formation was only about 2-fold higher than in control strains. The relatively small difference between normal and MSUD fibroblastsin vitro as opposed to the striking differences between healthy subjects and MSUD patientsin vivo are discussed with respect to the significance of physiological mechanisms participating in the formation and degradation ofl-alloisoleucine in man.

Two families with medium-chain acyl-CoA dehydrogenase (MCAD) deficiency due to compound heterozygosity are described. All patients have a 13 bp insertion in exon 11 of one allele at the MCAD gene locus. In the other allele patients in one of the families harbour the prevalent G985 mutation, and the other family possess an unidentified mutation causing reduced levels of MCAD mRNA. We demonstrate that the disease in these families is inherited as an autosomal recessive trait. Individuals heterozygous for the mutations show heterozygous/control levels of ß-oxidation activities in cultured fibroblasts (9.1–16.3 pmol/min per mg protein; control 10–17 pmol/min per mg protein), and in the excretion of the ‘ß-oxidation metabolites’, hexanoylglycine (<2 µmol/mmol creatinine), suberylglycine (<2 µmol/mmol creatinine) and phenylpropionylglycine (<2 µmol/mmol creatinine). This shows that there is no ‘negative dominance’ from the mutant monomeric protein onto the normal ones, in accordance with the finding of low levels of MCAD mRNA from the allele harbouring the 13 bp insertion as well as the allele with the unidentified mutation, and the low steady-state level of enzyme protein expressed from the G985-bearing allele. In the family possessing the G985 and the 13 bp insertion mutations, two asymptomatic compound heterozygous individuals were detected. They exhibited elevated excretion of hexanoylglycine (5–15 µmol/mmol creatinine) and suberylglycine (4–13 µmol/mmol creatinine), together with ß-oxidation activity in fibroblasts in the homozygous range (2.9 pmol/min per mg protein), showing a lack of correlation between the genotype, some biochemical parameters and the clinical phenotype.

l-Alloisoleucine formation froml-isoleucine was studiedin vitroandin vivo. When a healthy subject was loaded withl-isoleucine, plasma levels ofl-isoleucine and 3-methyl-2-oxopentanoate (KMV), as well asl-alloisoleucine, increased. Peak values were reached successively and were in the orderl-isoleucine > > KMV > >l-alloisoleucine. Metabolic clearance ofl-isoleucine and KMV was rapid; clearance ofl-alloisoleucine was considerably delayed. When human skin fibroblast cultures were challenged withl-isoleucine, KMV accumulated at a gradually decreased rate, whereasl-alloisoleucine accumulated at a gradually accelerated rate. KMV andl-alloisoleucine formation were related and depended on thel-isoleucine concentration applied. In cell lines derived from MSUD patients (classical form), metabolite formation was only about 2-fold higher than in control strains. The relatively small difference between normal and MSUD fibroblastsin vitroas opposed to the striking differences between healthy subjects and MSUD patientsin vivoare discussed with respect to the significance of physiological mechanisms participating in the formation and degradation ofl-alloisoleucine in man.