Methylmalonate semialdehyde

Unlike the end products of purine catabolism, those of pyrimidine catabolism are highly water-soluble COj, NH3, P-alanine, and P-aminoisobutyrate (Figure 34-9). Excretion of P-aminoisobutyrate increases in leukemia and severe x-ray radiation exposure due to increased destruction of DNA. However, many persons of Chinese or Japanese ancestry routinely excrete P-aminoisobutyrate. Humans probably transaminate P-aminoisobutyrate to methylmalonate semialdehyde, which then forms succinyl-CoA (Figure 19-2). 

Figure 10.8 A summary of the reactions involved in the degradation of nucleic acid, nucleotides, nucleosides and purine and pyn midine bases. Nucleic add is hydrolysed by nucleases to form nucleotides, which are hydrolysed to nucleosides. The latter are split into ribose 1-phosphate and a base. The purine bases are converted to uric acid and ammonia. Uric acid is excreted. The pyrimidine bases are converted to 3-carbon intermediates (malo-nate semialdehyde and methylmalonate semialdehyde). The nitrogen is released as ammonia or used to convert oxoglutarate to glutamate.

Serum carnosinase activity is readily measured as a marker for carnosinosis, homocarnosinosis and in instances of jS-alanine elevation in physiological fluids [7]. For quantification, carnosine is incubated with sera samples, and the histidine liberated in the reaction is quantified as the fluorescent o-phthalaldehyde derivative [19]. To estimate the activity of methylmalonate semialdehyde dehydrogenase (direct enzyme determination methods have not been reported), fibroblast extracts are incubated with l-14C-/ -alanine and trapping of 14C02. 

Fig. 6. Pathway leading from valine to methylmalonic acid. Metliylmalonic acid could arise through direct oxidation of methylmalonic semialdehyde, as shown in the scheme, or through deacylation of methylmalonyl-CoA, which would represent an intermediate (not shown on the scheme) between methylmalonic semialdehyde and methylmalonic acid.

The best-studied example of a CoA-dependent nonphosphorylating ALDH is the methylmalonate-semi-aldehyde dehydrogenase, which has been isolated from both mammalian and bacterial sources. This enzyme transforms malonate semialdehyde and methylmalonate semialdehyde into acetyl-CoA and propionyl-CoA, respectively, through an oxidation reaction as described above, followed by a decarboxylation reminiscent of other (3-keto acids. Mechanistic studies of the B. sukilis enzyme have shown that it is activated by NAD" " binding, that it exhibits half-of-sites reactivity (only two moles of NADH forms per tetrameric protein unit) and that the decarboxylation reaction occurs after formation of the acyl-enzyme intermediate. Acyl transfer from the enzyme to CoA completes the reaction. 

The transamination of P-aminoisobutyrate to form methylmalonate semialdehyde requires pyridoxal phosphate as a cofactor. This reaction is similar to the conversion of ornithine to glutamate y-semialdehyde. Then NAD+ serves as an electron acceptor for the oxidation of methylmalonate semialdehyde to methylmalonate. The conversion of methylmalonate to methylmalonyl CoA requires coenzyme A. The final reaction, in which methylmalonyl CoA is converted to succinyl CoA, is catalyzed by methylmalonyl CoA mutase, an enzyme that contains a derivative of vitamin B12 as its coenzyme. 

Hydroxisobutyrate is oxidized by a DPN-specific dehydrogenase TPN is not attacked and the enzyme does not attack any known analogs of either 8-hydroxyisobutyrate or the oxidation product, methylmalonate semialdehyde. The further degradation of the branched chain has not... 

Evidence for the mechanism of the conversion step of isobutyric to propionic acid (reaction 3, Fig. 4) was sought by Atchley on the premise that any compound which is an intermediate between isobutyrate and propionate should be completely oxidized by the fatty acid oxidizing system of liver. Using as a guide in the selection of compounds the 8-oxidation theory of fatty acids, he found that 8-hydroxyisobutyrate and methylacrylate were completely oxidized. Methylacrylic acid could be expected to give rise to methylmalonic semialdehyde, which, on structural grounds,... 

Methylmalonate semialdehyde dehydrogenase deficiency [8] has been described in a single patient. This patient, a boy, came to attention because of an elevated concentration of methionine on routine neonatal screening. The value exceeded 1000 imol/l. By 4 years of age he had developed normally. A valine load was followed by an increase in 3-hydroxyisobutyric acid excretion. Incubation of fibroblasts from the patient with 2-valine or p-[1- C]-alanine led to no production of C02 from valine and very little from )ff-alanine in contrast to control cells. 

Examination of plasma and urine revealed elevated quantities of jff-ala-nine, 3-hydroxypropionic acid, (R)- and (S)-3-aminoisobutyric acid, (R)-and (S)-3-hydroxyisobutyric acid and (S)-2-hydroxymethylbutyric acid. Direct enzymatic assay of methylmalonate semialdehyde dehydrogenase is unavailable revealed homozygosity for DNA analysis 1336 G>A transversion which substituted an arginine for a highly conserved glycine at amino acid residue 446. 

The site of the defect in methylmalonic semialdehyde dehydrogenase deficiency. 

Table 7.5. 3-Hydroxyisobutyric acidemia due to methylmalonate semialdehyde dehydrogenase deficiency...

Table 7.6. 3-Hydroxyisobutyric acidemia with normal methylmalonate semialdehyde dehydrogenase ...

S-Aminoisobutyric acid a-ketoglutaric acid methylmalonic semialdehyde H- glutamic acid SS4... 

The next product in the reaction sequence, methylmalonate semialdehyde, cannot be formed from the CoA thiol ester of hydroxyisobutyrate and requires free /3-hydroxy isobutyrate (87). Free /3-hydroxyisobutyrate and DPN react rapidly in the presence of the enzyme. Proof of the formation of the methylmalonate semialdehyde was obtained by cariying out the incubation in the presence of hydroxy lamine and converting the product to the methylmalonic semialdehyde phenylhydrazone. 

The optimum activity of this enzyme is at about pH 9.0 the Km value for /3-hydroxyisobutyrate was.estimated to be 1.2 X 10 M and for DPN, 3.3 X 10 M. The equilibrium constant, [(methylsemialdehyde)(DPNH) (H+)/(/3-hydroxyisobutyrate)(DPN+)l, was calculated to be 3 X 10 This shows that the equilibrium position markedly favors the reduction of the methylmalonate semialdehyde to the hydroxy compound. 

Methylmalonate semialdehyde has been shown to be transaminated to form /3-aminoisobutyrate, as well as being decarboxylated to propionyl aldehyde (89). Previously it had been determined that this compound was formed in the metabolism of dihydroth3mine (98, 99). Kupiecki and Coon (89) prepared a transaminase from pig kidney which catalyzes the transamination of methylmalonate semialdehyde with glutamic acid as the amino group donor to /3-aminoisobutyric acid according to Eq. (6). 

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