Molecular organization of peroxisomal enzymes: protein-protein interactions in the membrane and in the matrix.
The beta-oxidation of fatty acids in peroxisomes produces hydrogen peroxide (H2O2), a toxic metabolite, as a bi-product. Fatty acids beta-oxidation activity is deficient in X-linked adrenoleukodystrophy (X-ALD) because of mutation in ALD-gene resulting in loss of very long chain acyl-CoA synthetase (VLCS) activity. It is also affected in disease with ... catalase negative peroxisomes as a result of inactivation by H2O2. Therefore, the following studies were undertaken to delineate the molecular interactions between both the ALD-gene product (adrenoleukodystrophy protein, ALDP) and VLCS as well as H2O2 degrading enzyme catalase and proteins of peroxisomal beta-oxidation. Studies using a yeast two hybrid system and surface plasmon resonance techniques indicate that ALDP, a peroxisomal membrane protein, physically interacts with VLCS. Loss of these interactions in X-ALD cells may result in a deficiency in VLCS activity. The yeast two-hybrid system studies also indicated that catalase physically interacts with L-bifunctional enzyme (L-BFE). Interactions between catalase and L-BFE were further supported by affinity purification, using a catalase-linked resin. The affinity bound 74-kDa protein, was identified as L-BFE by Western blot with specific antibodies and by proteomic analysis. Additional support for their interaction comes from immunoprecipitation of L-BFE with antibodies against catalase as a catalase- L-BFE complex. siRNA for L-BFE decreased the specific activity and protein levels of catalase without changing its subcellular distribution. These observations indicate that L-BFE might help in oligomerization and possibly in the localization of catalase at the site of H2O2 production in the peroxisomal beta-oxidation pathway.
Mesh Terms:
3-Hydroxyacyl CoA Dehydrogenases, Animals, Binding Sites, Biological Markers, Catalase, Coenzyme A Ligases, Cytosol, Enoyl-CoA Hydratase, Extracellular Matrix, Humans, Intracellular Membranes, Isomerases, Liver, Molecular Weight, Multienzyme Complexes, Oxidation-Reduction, Peptide Fragments, Peroxisomes, Protein Binding, Proteome, Proteomics, RNA Interference, RNA, Messenger, Rats, Rats, Sprague-Dawley, Sensitivity and Specificity, Subcellular Fractions
3-Hydroxyacyl CoA Dehydrogenases, Animals, Binding Sites, Biological Markers, Catalase, Coenzyme A Ligases, Cytosol, Enoyl-CoA Hydratase, Extracellular Matrix, Humans, Intracellular Membranes, Isomerases, Liver, Molecular Weight, Multienzyme Complexes, Oxidation-Reduction, Peptide Fragments, Peroxisomes, Protein Binding, Proteome, Proteomics, RNA Interference, RNA, Messenger, Rats, Rats, Sprague-Dawley, Sensitivity and Specificity, Subcellular Fractions
Arch. Biochem. Biophys.
Date: Jul. 15, 2006
PubMed ID: 16781659
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