論文 - 詳細
| RRC ID | 29143 |
|---|---|
| 著者 | Sumida T, Sueyoshi N, Ito M. |
| タイトル | Molecular cloning and characterization of a novel glucocerebrosidase of Paenibacillus sp. TS12. |
| ジャーナル | J Biochem |
| Abstract |
We report here the molecular cloning and characterization of a glucocerebrosidase [EC 3.2.1.45] from Paenibacillus sp. TS12. The open reading frame of the glucocerebrosidase gene consisted of 2,493 bp nucleotides and encoded 831 amino acid residues. The enzyme exhibited no sequence similarity with a classical glucocerebrosidase belonging to glycoside hydrolase (GH) family 30, but rather showed significant similarity with GH family 3 beta-glucosidases from Clostridium thermocellum, Ruminococcus albus, and Aspergillus aculeateus. The recombinant enzyme, expressed in Escherichia coli BL21(DE3)pLysS, had a molecular weight of 90.7 kDa and hydrolyzed NBD-labeled glucosylceramide, but not galactosylceramide, GM1a or sphingomyelin. The enzyme was most active at pH 6.5, and its apparent Km and Vmax values for NBD-labeled glucosylceramide and p-nitrophenyl-beta-glucopyranoside were 223 microM and 1.60 micromol/min/mg of protein, and 593 microM and 112 micromol/min/mg of protein, respectively. Site-directed mutagenesis indicated that Asp-223 is an essential amino acid for the catalytic reaction and possibly functions a catalytic nucleophile, as in GH family 3 beta-glucosidases. This is the first report of the molecular cloning and characterization of a glucocerebrosidase from a procaryote. |
| 巻・号 | 132(2) |
| ページ | 237-43 |
| 公開日 | 2002-8-1 |
| DOI | 10.1093/oxfordjournals.jbchem.a003216 |
| PMID | 12153721 |
| MeSH | Amino Acid Sequence Animals Bacteria / enzymology* Bacteria / genetics Base Sequence Cloning, Molecular Glucosylceramidase / chemistry Glucosylceramidase / genetics Glucosylceramidase / metabolism* Glucosylceramides / metabolism Humans Molecular Sequence Data Molecular Weight Mutation Open Reading Frames Oxadiazoles / metabolism Recombinant Proteins / genetics Recombinant Proteins / metabolism Sequence Alignment Substrate Specificity |
| IF | 2.476 |
| 引用数 | 12 |
| WOS 分野 | BIOCHEMISTRY & MOLECULAR BIOLOGY |
| オルトメトリクス指標 |
オルトメトリクス指標項目
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| 最多言及媒体 | Patent(IFI CLAIMS) |
| 各媒体での言及数の合計 | 1 |
| 過去6か月間でのオルトメトリクス指標の変動値 | 0.0 |
| リソース情報 | |
| 遺伝子材料 | Glc4 (RDB03002) pETGlc4 (RDB03003) |