Copurification of dihydroxyacetone-phosphate acyl-transferase and other peroxisomal proteins from liver of fenofibrate-treated rats
| Autor: | C. Causeret, Marc Bentejac, Maurice Bugaut, B. Teubner, Sabrina Albet |
|---|---|
| Rok vydání: | 1997 |
| Předmět: |
Male
Molecular Sequence Data Biochemistry Microbodies Copurification chemistry.chemical_compound Fenofibrate Protein purification Animals Amino Acid Sequence Rats Wistar Peptide sequence Dihydroxyacetone phosphate chemistry.chemical_classification Oxidase test Chromatofocusing Membrane Proteins General Medicine Peroxisome Molecular biology Rats Enzyme chemistry Liver Solubility Sequence Analysis Acyltransferases |
| Zdroj: | Biochimie. 79(7) |
| ISSN: | 0300-9084 |
| Popis: | Dihydroxyacetone-phosphate acyl-transferase (DHAP-AT), a peroxisomal membrane-bound enzyme that catalyzes the first step of ether-glycerolipid synthesis, was purified from liver of rats treated with fenofibrate, a peroxisome proliferator. The protocol first included isolation of peroxisomes, their purification through a discontinuous gradient and solubilization of membranes in CHAPS. DHAP-AT was further purified by four chromatographic steps, namely low-pressure size-exclusion, cation-exchange, hydroxylapatite and chromatofocusing. The chromatofocusing step led to a 4000-fold increase in the specific activity of DHAP-AT with respect to the liver homogenate with a yield of about 0.2%. Trypsin digestion of a 64-kDa protein band upon SDS-PAGE resulted in a peptide sequence unknown in databases. A corresponding degenerated oligonucleotide was used as a probe in Northern blotting, and a transcript of 3.3 kb was detected in some rat tissues. Moreover, the overall procedure allowed co-purification of four major peroxisomal enzymes: urate-oxidase, catalase, multifunctional enzyme and palmitoyl-CoA oxidase, respectively. |
| Databáze: | OpenAIRE |
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