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PMID:18584171
| Citation |
Kanamasa, S, Dwiarti, L, Okabe, M and Park, EY (2008) Cloning and functional characterization of the cis-aconitic acid decarboxylase (CAD) gene from Aspergillus terreus. Appl. Microbiol. Biotechnol. 80:223-9 |
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| Abstract |
A filamentous fungus Aspergillus terreus produces itaconic acid, which is predicted to be derived from cis-aconitic acid via catalysis by cis-aconitic acid decarboxylase (CAD) in the carbon metabolism of the fungus. To clarify the enzyme's function and a pathway for itaconic acid biosynthesis, we cloned a novel gene encoding the enzyme. The open reading frame of this gene (CAD1) consists of 1,529 bp encoding 490 amino acids and is interrupted by a single intron. Among the identified proteins in the database, the primary structure of the protein encoded by CAD1 shared high identity with the MmgE/PrpD family of proteins, including a number of 2-methylcitrate dehydratases of bacteria. The cloned gene excluding an intron was introduced into the expression plasmid pAUR-CAD1 controlled by the ADH1 promoter. The CAD activity in Saccharomyces cerevisiae was confirmed by directly detecting itaconic acid as a product from cis-aconitic acid as a substrate. This result reveals for the first time that this gene encodes CAD, which is essential for itaconic acid production in A. terreus. |
| Links |
PubMed Online version:10.1007/s00253-008-1523-1 |
| Keywords |
Aconitic Acid/metabolism; Amino Acid Sequence; Aspergillus/chemistry; Aspergillus/enzymology; Aspergillus/genetics; Base Sequence; Carboxy-Lyases/chemistry; Carboxy-Lyases/genetics; Carboxy-Lyases/isolation & purification; Carboxy-Lyases/metabolism; Cloning, Molecular; Fungal Proteins/chemistry; Fungal Proteins/genetics; Fungal Proteins/isolation & purification; Fungal Proteins/metabolism; Gene Expression; Molecular Sequence Data; Open Reading Frames; Saccharomyces cerevisiae/genetics; Saccharomyces cerevisiae/metabolism; Succinates/metabolism |
| edit table |
Significance
Annotations
| Gene product | Qualifier | GO Term | Evidence Code | with/from | Aspect | Extension | Notes | Status |
|---|---|---|---|---|---|---|---|---|
| GO:0047613: aconitate decarboxylase activity |
ECO:0000315: |
F |
The CAD1 gene was cloned from and then expressed in S. cerevisiae where activity was verified. Fig. 3 shows how the recombinant activity (measured by direct detection of itaconic acid product) was detected by HPLC. |
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