Analysis of insecticidal proteins from Bacillus thuringiensis and recombinant Escherichia coli by capillary electrokinetic chromatography.
Bacillus thuringiensis and recombinant Escherichia coli proteinaceous protoxins were subject to proteolysis and analyzed by capillary electrokinetic chromatography. Three resulting toxins (65 kDa) were baseline-resolved within 22 min using a 10 mM borate, pH 11 separation buffer consisting of 25 mM sodium dodecyl sulfate (SDS) and 30 mM phytic acid. The toxins displayed differential interactions with the SDS and phytic acid phases to effect their separation. The ion-pairing interaction between the analyte and phytic acid was also useful in preventing adsorption to the capillary walls and thus enhanced separation resolution and efficiency. The use of electrokinetic chromatography allows achievement of the separation in a significantly shorter time than conventional high-performance liquid chromatography (HPLC) using a diethylaminoethyl (DEAE) weak-anion exchanger.
Biotechnology Research Institute, National Research Council Canada, Montreal, Quebec, Canada. john.luong@cnrc-nrc.gc.ca
John H T
JH
Luong
Keith B
KB
Male
Alberto
A
Mazza
Luke
L
Masson
Roland
R
Brousseau
D001413:Bacillus thuringiensis / Q000235:genetics*; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000032:analysis*; D001427:Bacterial Toxins / Q000032:analysis*; D020374:Chromatography, Micellar Electrokinetic Capillary / Q000379:methods*; D004731:Endotoxins / Q000032:analysis*; D004926:Escherichia coli / Q000235:genetics*; D006460:Hemolysin Proteins; D007536:Isomerism; D010833:Phytic Acid / Q000737:chemistry; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D012967:Sodium Dodecyl Sulfate; D010833:Phytic Acid