Complete cDNA cloning and prokaryotic expression of chitin deacetylase CDA2 from Mucor racemosus
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Key Words: chitin deacetylase  cda2  cDNA cloning  Mucor racemosus  prokaryotic expression
Author NameAffiliation
JIANG Xia-yun 上海海洋大学 食品学院 
YUAN Xiang-nan 上海海洋大学 农业部水产种质资源与利用重点开放实验室 
WEI Fu-wei 上海海洋大学 食品学院 
ZHOU Pei-gen 上海海洋大学 食品学院 
ZOU Shu-ming 上海海洋大学 农业部水产种质资源与利用重点开放实验室 
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      The complete cDNA (GenBank accession number DQ678929) and its corresponding gene (EF468349) of chitin deacetylase CDA2 from Mucor racemosus mycelium had been cloned by rapid amplification cDNA end (RACE) with specific degenerate primers. In contrast to previously obtained gene cda1, cda2 contained no intron sequence. It consisted of 1 378 bp nucleotides, comprising 23-bp 5’ untranslated region (UTR), 1 254 bp open reading frame (ORF) and 101-bp 3’ UTR. The ORF encoded 418 amino acid (a.a.) residues including a 21 a.a. N-terminal signal peptide and a conserved polysaccharide deacetylase domain were located in an area of 150-272 a.a. residues. The results from subsequent construction of expressional vector pET28a-cda2 and prokaryotic expression revealed that molecular weight of recombinant protein CDA2 was about 46 ku and it was mainly found in inclusion bodies. The purified CDA2 showed chitin deacetylating activities. This work is necessary for further structural and functional exploration in chitin deacetylase CDA2 from M. racemosus.