Propionyl coenzyme A carboxylase

The method described is suitable for the assay of four biotin-containing carboxylases pyruvate carboxylase, acetyl-coenzyme A carboxylase, propionyl-coenzyme A carboxylase, and 3-methylcrotonyl-coenzyme A carboxylase. The assays do not require radioisotopes and are suitable for use in clinical laboratories. 

Substrates and products are separated by reversed-phase chromatography at 45°C on a Nucleosil Qs column (4.6 mm X 250 mm). For assay of acetyl-coenzyme A carboxylase, propionyl-coenzyme A carboxylase, and 3-methylcrotonyl-coenzyme A carboxylase, a linear gradient from solvent A (0.1 M sodium phosphate buffer, pH 2.1) to solvent B (methanol-solvent A, 80 20, v/v) was applied in 15 minutes at a flow rate of 1.5 mL/min. Quantitation was based on the absorbance of the product (malonyl-CoA, methylmalonyl-CoA, and 3-methylglutaconyl-CoA, respectively) at 260 nm. For assay of pyruvate carboxylase, pyruvate was separated by isocratic elution using 0.1 M sodium phosphate buffer (pH 2.1) containing 0.1 M sodium sulfate. Quantitation was based on the disappearance of pyruvate as followed at 210 nm. 

Biotin serves as the prosthetic group of several enzymes that catalyse the transfer of carbon dioxide from one substrate to another. In animals there are three biotin-dependent enzymes of particular importance pyruvate carboxylase (carbohydrate synthesis from lactate), acetyl coenzyme A carboxylase (fatty acid synthesis) and propionyl coenzyme A carboxylase (the pathway of conversion of propionate to succinyl-CoA). The specific role of these enzymes in metabolism is discussed in Chapter 9. 

Harris, D.J., Yang, B.I., Wolf, B. and Snodgrass, P.J. (1980), Dysautonomia in an infant with secondary hyperammonemia due to propionyl Coenzyme-A carboxylase deficiency. Pediatrics, 65,107. 

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