Amino acid substitution on Beta1 and alphaF of Cyt2Aa2 affects molecular interaction of protoxin.
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 Beta1 and C-terminal alphaF 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 Beta1 has a critical effect on dimer formation and structural stability against proteases. In addition, amino acids N230 and I233-F237 around the C-terminus alphaF demonstrated a crucial role in protecting the protoxin from proteolytic digestion. These results suggested that Beta1 and alphaF 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.
Thammachat
Siriya
S
Institute of Molecular Biosciences, Mahidol University, Salaya, Nakhon Pathom, Thailand.
Pungtanom
Nuanwan
N
Kidsanguan
Somruathai
S
Pathaichindachote
Wanwarang
W
Promdonkoy
Boonhiang
B
Krittanai
Chartchai
C
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001427:Bacterial Toxins / Q000737:chemistry* / Q000235:genetics; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D013050:Spectrometry, Fluorescence
D001427:Bacterial Toxins