Alanine-162 of Bacillus thuringiensis Cyt2Aa2 toxin is essential for membrane binding and oligomerisation

作者：Saitonuang Pajaree; Promdonkoy Boonhiang; Kubera Anchanee^*

来源：Biocontrol Science and Technology, 2016, 26(7): 928-937.

DOI：10.1080/09583157.2016.1168776

• Cyt2aa2
• Bacillus thuringiensis
• membranebinding residues
• Aedes aegypti
• site-directed mutagenesis

Cyt2Aa2 is a cytolytic toxin produced by Bacillus thuringiensis subsp. darmstadiensis. It is specifically toxic to dipteran larvae in vivo and is also active against several cell types, such as erythrocytes. The active toxin is proposed to bind to the cell membrane, and membrane pore formation by toxin oligomerisation leads to cell lysis. This study aimed to characterise the role of residues (I139, S159, L160, S161, A162, D209 and V215) potentially involved in the membrane binding of Cyt2Aa2. All mutants, except I139A and V215A, showed similar characteristics to the wild-type toxin after proteinase K cleavage. Three mutants, S159A, L160A and S161A, showed high haemolytic activity but low toxicity against Aedes aegypti. Membrane interaction assays showed that these mutants could bind to rat red blood cells (rRBCs) and oligomerise. The mutant D209N had no haemolytic activity but was still mildly toxic to A. aegypti. The mutant A162V could not lyse rRBCs, even at high concentrations, and showed no toxicity against A. aegypti. Our data suggest that alanine 162 of the Cyt2Aa2 toxin is involved in membrane binding and oligomerisation. Substitution of this amino acid altered the conformation of the toxin and affected its biological activity.

• 出版日期2016-7