Back to Search
Start Over
Highly heterogeneous nature of delta-aminolevulinate dehydratase (ALAD) deficiencies in ALAD porphyria.
- Source :
-
Blood [Blood] 2001 May 15; Vol. 97 (10), pp. 2972-8. - Publication Year :
- 2001
-
Abstract
- The properties of 9 delta-aminolevulinate dehydratase (ALAD) mutants from patients with ALAD porphyria (ADP) were examined by bacterial expression of their complementary DNAs and by enzymologic and immunologic assays. ALADs were expressed as glutathione-S-transferase (GST) fusion proteins in Escherichia coli and purified by glutathione-affinity column chromatography. The GST-ALAD fusion proteins were recognized by anti-ALAD antibodies and were enzymatically active as ALAD. The enzymatic activities of 3 ALAD mutants, K59N, A274T, and V153M, were 69.9%, 19.3%, and 41.0% of that of the wild-type ALAD, respectively, whereas 6 mutants, G133R, K59N/G133R, F12L, R240W, V275M, and delTC, showed little activity (< 8%). These variations generally reflect the phenotype of ALAD in vivo in patients with ADP and indicate that GST-ALAD fusion protein is indeed useful for predicting of the phenotype of ALAD mutants. The location of F12L mutation in the enzyme's molecular structure indicates that its disturbance of the quaternary contact of the ALAD dimer appears to have a significant influence on the enzymatic activity. Mouse monoclonal antibodies to human ALAD were developed that specifically recognized a carboxy terminal portion of ALAD, or other regions in the enzyme. This study represents the first complete analysis of 9 mutants of ALAD identified in ADP and indicates the highly heterogeneous nature of mutations in this disorder.
- Subjects :
- Adolescent
Animals
Antibodies, Monoclonal immunology
Antibody Specificity
Escherichia coli genetics
Female
Gene Expression
Glutathione Transferase genetics
Humans
Immunoblotting
Infant, Newborn
Male
Mice
Mice, Inbred BALB C
Middle Aged
Phenotype
Porphobilinogen Synthase metabolism
Porphyrias genetics
Recombinant Fusion Proteins analysis
Recombinant Fusion Proteins isolation & purification
Recombinant Fusion Proteins metabolism
Mutation
Porphobilinogen Synthase deficiency
Porphobilinogen Synthase genetics
Porphyrias enzymology
Subjects
Details
- Language :
- English
- ISSN :
- 0006-4971
- Volume :
- 97
- Issue :
- 10
- Database :
- MEDLINE
- Journal :
- Blood
- Publication Type :
- Academic Journal
- Accession number :
- 11342419
- Full Text :
- https://doi.org/10.1182/blood.v97.10.2972