[N-terminal analysis and antibody preparation of insecticidal protein form Pseudomonas pseudoalcaligenes]

The insecticidal protein from Pseudomonas pseudoaligenes was and exotoxin which had toxicity on locusts. In order to elucidate its molecular properties an amino acid sequence, the insecticidal protein was purified from the culture supernatant by ultrafiltration, ion-exchange chromatography and gel filtration, and showed a single band on SDS-PAGE. Analysis of the purified insecticidal protein dentified N-terminal sequence of ten amino acid residues. Its polyclonal antibody was also obtained by immunizing rabbit with the insecticidal protein recovered form SDS-PAGE gel. The antibody titer determined by ELISA method was 1:12,800, indicating that it has high reactivity. Western blot analysis revealed that the antibody was spectific to 26 kD insecticidal protein, and did not cross-react with other proteins produced by the bacterium, suggesting that a specific antibody with high titer was obtained and could be used for further investigations of the gene cloning and expression of insecticidal protein.