Cloning and Functional Identification of Delta5 Fatty Acid Desaturase Gene and Its 5′-Upstream Region from Marine Fungus Thraustochytrium sp. FJN-10
| Autor: | Xiao-Feng Xia, Xiao-Feng Chen, Ruo-Yu Mao, Baoyu Tian, Jianzhong Huang, Aiqun Yu, Xianzhang Jiang |
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| Rok vydání: | 2010 |
| Předmět: |
Fatty Acid Desaturases
Reading Frames Molecular Sequence Data Saccharomyces cerevisiae Reading frame Applied Microbiology and Biotechnology Fungal Proteins Amino Acid Sequence Cloning Molecular Peptide sequence Gene chemistry.chemical_classification Genetics Base Sequence biology Fungi Nucleic acid sequence biology.organism_classification Amino acid Open reading frame Fatty acid desaturase Biochemistry chemistry Fatty Acids Unsaturated biology.protein Stramenopiles |
| Zdroj: | Marine Biotechnology. 13:12-21 |
| ISSN: | 1436-2236 1436-2228 |
| DOI: | 10.1007/s10126-010-9262-6 |
| Popis: | A gene encoding delta5 fatty acid desaturase (fad5) was cloned from marine fungus Thraustochytrium sp. FJN-10, a species capable of producing docosahexaenoic acid. The open reading frame of fad5 was 1,320 bp and encoded a protein comprising 439 amino acids. Expression of the fad5 in Saccharomyces cerevisiae INVSC1 revealed that FAD5 is able to introduce a double bond at position 5 of the dihomo-γ-linolenic acid (20:3 Δ(8,11,14)), resulting in arachidonic acid (20:4 Δ(5,8,11,14)) with a conversion rate of 56.40% which is the highest among engineering yeasts reported so far. The 5'-upstream region of fad5 was cloned by LA-PCR and analyzed. Phylogenetic analysis of this sequence with the 5'-upstream region of other delta5 desaturases showed that the 5'-upstream region of fad5 from Thraustochytrium share the smallest evolution distance with human and rhesus. Computational analysis of the nucleotide sequence of the 5'-upstream region of fad5 has revealed several basic transcriptional elements including five TATA boxes, three CCAAT boxes, 12 GC boxes, and several putative target-binding sites for transcription factors such as HSF, CAP, and ADR1. Preliminary functional analysis of this promoter in S. cerevisiae shows that the 5'-upstream region of fad5 could drive the expression of green fluorescent protein. |
| Databáze: | OpenAIRE |
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