[Chitinase B from Bacillus thuringiensis enhancing potential and its antagonism and insecticidal].
OBJECTIVE
We studied the characteristics of chitinase B (ChiB) from Bacillus thuringiensis subsp. colmeri 15A3 (Bt 15A3) and evaluated its antagonism and insecticidal potential.


METHODS
ChiB was expressed in Escherichia coli. Expression product was purified by ammonium sulfate precipitation, dialysis and Sephadex G-200 gel filtration chromatography. The molecular mass of ChiB was directly estimated by zymogram after sodium dodecyl sulfate-polyacryamide gel electroresis (SDS-PAGE). The proteins with different molecular weight were identified by mass spectrum (MS) and analyzed through bioinformatics. We studied some metal ions' impacts on chitinase activity; optimal temperature, pH and its stability at different temperature and pH. We also studied the inhibition effects on sporangia germination of fungi and the synergistic effects of ChiB on larvicidal activity.


RESULTS
The molecular mass of ChiB was estimated as 70 kDa by zymogram. The protein with 64 kDa was the product of C-terminal degradation of ChiB in E. coli. Chitinase activity was improved by Fe3+ and inhibited by Zn2+ and Ag2+. The optimal temperature for ChiB was 60 C and the optimal pH was 5.0. The enzyme was quite stable at temperature below 60 degrees C and pH between 4.0 and 8.0. ChiB inhibited sporangia germination and the IC50 (50% inhibited concentration) was about 4 microg/mL. Moreover, the bioassays showed that ChiB could reduce the LC50 (50% lethal concentration) of the crystal protein of Bt. against Spodoptera exigua and Helicoverpa armigera larvae by approximately 26% and 30%.


CONCLUSIONS
ChiB was not only stable, but also with good antagonism and insecticidal activity.
Liu
Dong
D
Department of Microbiology, Nankai University, Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Tianjin 300071, China. liudong204@mail.nankai.edu.cn
Chen
Yuehua
Y
Cai
Jun
J
Xiao
Liang
L
Liu
Chuan
C
D000818:Animals; D001413:Bacillus thuringiensis / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology*; D002688:Chitinases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology*; D004795:Enzyme Stability; D005658:Fungi / Q000187:drug effects* / Q000502:physiology; D007306:Insecticides / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D008970:Molecular Weight; D009036:Moths / Q000187:drug effects*
D001426:Bacterial Proteins; D007306:Insecticides; D002688:Chitinases