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http://dx.doi.org/10.5483/BMBRep.2010.43.6.427

Amino acid substitution on β and α of Cyt2Aa2 affects molecular interaction of protoxin  

Thammachat, Siriya (Institute of Molecular Biosciences, Mahidol University, Salaya Campus)
Pungtanom, Nuanwan (Institute of Molecular Biosciences, Mahidol University, Salaya Campus)
Kidsanguan, Somruathai (Institute of Molecular Biosciences, Mahidol University, Salaya Campus)
Pathaichindachote, Wanwarang (National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency)
Promdonkoy, Boonhiang (National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency)
Krittanai, Chartchai (Institute of Molecular Biosciences, Mahidol University, Salaya Campus)
Publication Information
BMB Reports / v.43, no.6, 2010 , pp. 427-431 More about this Journal
Abstract
Cyt2Aa2 is a mosquito-larvicidal protein produced as a 29 kDa crystalline protoxin from Bacillus thuringiensis subsp. darmstadiensis. To become an active toxin, proteolytic processing is required to remove amino acids from its N- and C-termini. This study aims to investigate the functional role of amino acid residues on the N-terminal ${\beta}1$ and C-terminal ${\alpha}F$ of Cyt2Aa2 protoxin. Mutant protoxins were constructed, characterized and compared to the wild type Cyt2Aa2. Protein expression data and SDS-PAGE analysis revealed that substitution at leucine-33 (L33) of ${\beta}1$ has a critical effect on dimer formation and structural stability against proteases. In addition, amino acids N230 and I233-F237 around the C-terminus ${\alpha}F$ demonstrated a crucial role in protecting the protoxin from proteolytic digestion. These results suggested that ${\beta}1$ and ${\alpha}F$ on the Nand C-terminal ends of Cyt2Aa2 protoxin play an important role in the molecular interaction and in maintaining the structural stability of the protoxin.
Keywords
Bacillus thuringiensis; Cytolytic toxin; Mutagenesis; Protein folding; Toxicity;
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