| RRC ID |
36623
|
| 著者 |
Razeto A, Mattiroli F, Bossi R, Coda A, Mattevi A.
|
| タイトル |
Identifying a recombinant alkyldihydroxyacetonephosphate synthase suited for crystallographic studies.
|
| ジャーナル |
Protein Expr Purif
|
| Abstract |
Alkyldihydroxyacetonephosphate is the building block for the biosynthesis of ether phospholipids, which are essential components of eukaryotic cell membranes and are involved in a variety of signaling processes. The metabolite is synthesized by alkyldihydroxyacetonephosphate synthase (ADPS), a peroxisomal flavoenzyme. Deficiency in ADPS activity causes rhizomelic chondrodysplasia punctata type 3, a very severe genetic disease. ADPS is unusual in that it uses a typical redox cofactor such as FAD to catalyze a non-redox reaction. With the goal of undertaking a structural investigation of the enzyme, we have characterized recombinant ADPS from different sources: Cavia porcellus, Drosophila melanogaster, Homo sapiens, Archaeoglobus fulgidus, and Dictyostelium discoideum. The protein from D. discoideum was found to be the best candidate for structural studies. We describe a protocol for expression and purification of large amounts of pure and stable enzyme in its holo (FAD-bound) form. A search of deletion mutants identified a protein variant that forms crystals diffracting up to 2A resolution.
|
| 巻・号 |
55(2)
|
| ページ |
343-51
|
| 公開日 |
2007-10-1
|
| DOI |
10.1016/j.pep.2007.05.012
|
| PII |
S1046-5928(07)00139-8
|
| PMID |
17601746
|
| MeSH |
Alkyl and Aryl Transferases / chemistry
Alkyl and Aryl Transferases / metabolism*
Amino Acid Sequence
Animals
Base Sequence
Crystallography, X-Ray
DNA Primers
Electrophoresis, Polyacrylamide Gel
Humans
Hydrolysis
Molecular Sequence Data
Recombinant Proteins / chemistry
Recombinant Proteins / metabolism
Sequence Homology, Amino Acid
Spectrophotometry, Ultraviolet
|
| IF |
1.513
|
| 引用数 |
1
|
|
WOS 分野
|
BIOTECHNOLOGY & APPLIED MICROBIOLOGY
BIOCHEMICAL RESEARCH METHODS
BIOCHEMISTRY & MOLECULAR BIOLOGY
|
| リソース情報 |
| 細胞性粘菌 |
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