Evidence that the CryIA crystal protein from Bacillus thuringiensis is associated with DNA.
Toxin generated by activation of the Bacillus thuringiensis CryIA(c) crystal protein (protoxin) with bovine trypsin was separated into two components by anion-exchange chromatography. One component (T2) was DNA-associated toxin, and the other was the DNA-free toxin (T1). Only one major protoxin component was observed, and it was found to be associated with DNA. The DNA from the T2 toxin varied in size from 100 to 300 base pairs, whereas the crystal and the solubilized protoxin contained 20-kilobase DNA as the major DNA component. DNase treatment converted the T2 toxin to the DNA-free T1 toxin. In contrast, the DNA in the crystal and the solubilized protoxin was resistant to DNase digestion and was not dissociated from the protein by 1.5 M NaCl. The protoxin and DNA appeared to elute as a complex with a molecular mass of > 2 x 10(6) Da on gel-filtration chromatography. No toxin was generated from the protoxin with trypsin after extensive digestion of the protoxin with DNase or dissociation of the DNA by succinylation of the lysine residues. It is proposed that DNA binds to the COOH-terminal half of the crystal protein and is essential for maintaining the conformational integrity required for crystal formation and generation of toxin.
Bietlot
H P
HP
Department of Chemistry, University of Ottawa, Ontario, Canada.
Schernthaner
J P
JP
Milne
R E
RE
Clairmont
F R
FR
Bhella
R S
RS
Kaplan
H
H
D001413:Bacillus thuringiensis / Q000737:chemistry*; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry* / Q000302:isolation & purification; D001427:Bacterial Toxins / Q000737:chemistry* / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D004269:DNA, Bacterial / Q000737:chemistry* / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D004731:Endotoxins; D006460:Hemolysin Proteins; D008856:Microscopy, Fluorescence; D008970:Molecular Weight; D013056:Spectrophotometry, Ultraviolet; D014357:Trypsin
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004269:DNA, Bacterial; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D014357:Trypsin