Assay for l-pipecolate oxidase activity in human liver: detection of enzyme deficiency in hyperpipecolic acidaemia |
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Authors: | Vallabhaneni V. Rao Yung-Feng Chang |
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Affiliation: | Department of Biochemistry, University of Maryland, Dental School, Baltimore, MD, USA |
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Abstract: | A direct assay method is described for l-pipecolate oxidase. The assay uses NaHSO3 to trap the δ-semialdehyde (AAS) formed as a direct reaction product of l-pipecolate oxidase from l-[3H]pipecolic acid. The adduct so formed was separated from the substrate on Dowex 50 (H+) column. The product was identified as [3H]AAS by amino acid analysis after breaking down the adduct by boiling under acidic conditions. The assay is simpler and more specific than fluorometric methods; it is also more sensitive; requiring at most 16 μg of liver peroxisome-enriched protein per assay. We have used this assay procedure to detect l-pipecolate oxidase in skin fibroblasts obtained from a control subject and from patients of hyperpipecolic acidaemia and Zellweger syndrome and found that this enzyme activity is present in the control, but absent or decreased in the patients with the peroxisomal disorders. |
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Keywords: | Pipecolate oxidase Peroxisomal Zellweger Hyperpipecolic acidemia AAS Correspondence: Y.F. Chang, Department of Biochemistry, University of Maryland, Dental School, 666 West Baltimore Street, Baltimore, MD 21201, USA |
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