Graduating with research distinction in biochemistryThe insecticidal crystal proteins, known as Cry toxins, are naturally derived from Bacillus thuringiensis and provide an environmentally safe form of pest control. Though these proteins have been studied extensively, little is known regarding their specific mechanism of action and any conformational changes associated with this mechanism. In an effort to design a mutant Cry1Ab toxin with two individual lysine and cysteine residues for fluorophore ligation, this study focuses on the characterization of the structural effects of lysine mutagenesis leading to future development of such a toxin. In this study, the three lysines of active Cry1Ab toxin are mutated to alanine in different combinations, and the structural effects of these mutations are monitored by SDS-PAGE, bioassay, and circular dichroism wavelength scanning. Structural analyses have revealed that mutation of lysine 490 to alanine results in a toxin with increased protease sensitivity that makes isolation of pure, active toxin by the standardized procedure extremely difficult. Mutations of the other lysine residues are shown to have a lesser effect on toxin structure. The observations of this study contribute to data regarding potential sites on Cry1Ab for ligation of functional groups, and the structural and functional effects of mutagenesis at these sites.No embarg
