Maple Syrup Urine Disease: Coenzyme function and prenatal monitoring

A pregnancy at high risk for “cofactor resistant” Maple Syrup Urine Disease (MSUD) was monitored by quantitating valine, leucine, and isoleucine concentrations in maternal 24-hr urines, maternal plasma, and amniotic fluid. The fetal genotype was determined by assaying the conversion of radiolabeled branched-chain amino acids to 14CO 2 by intact cultured amniotic fluid cells. Although branched-chain amino acid concentrations in maternal fluids were similar to control values, cells cultured from the high-risk pregnancy produced 14CO 2 at one-half the rate of control cells. This finding suggested that the unborn 46 XY fetus was heterozygous for the MSUD gene. To test this hypothesis and to study normal and mutant branched-chain α-keto-acid dehydrogenase and co-factor interaction, a broken-cell system was developed that decarboxylated branched-chain α-keto-acids only when reconstituted with required cofactors. In control systems reduced coenzyme A (CoASH), β-nitotinamide adenine dinucleotide (NAD), thiamine pyrophosphate (TPP), and magnesium chloride (Mg 2+) stimulated 14CO 2 production from α-ketoisocaproic acid-I- 14C (KIC), α-ketoisovaleric acid-I- 14C (KIV), and L-α-keto-β-methylvaleric acid-I- 14C (KMV) by five to fifteen times base line over a 2-hr time course. TPP and Mg 2+ alone failed to increase either KIC or KIV decarboxylase, but reconstituted 30% of KMV decarboxylase. This “TPP-reconstituted” KMV decarboxylase was also present in mutant cells homozygous for this MSUD gene. KIV decarboxylase activity was essentially absent in the homozygous-affected line and partially impaired in heterozygous lines. Cells from the newborn male off-spring had partial impairment of KIV and KMV decarboxylase. These studies indicated that the MSUD mutation in this family resulted in the production of a defective subunit of the branched-chain α-keto-acid dehydrogenase complex that was common to all three branched-chain α-keto-acids, that a “TPP-reconstituted” KMV decarboxylase was present and under separate genetic control, and that a heterozygote for this MSUD gene was predicted before birth.