Expression and characterization of a recombinant Cry1Ac crystal protein with enhanced green fluorescent protein in acrystalliferous Bacillus thuringiensis.
OBJECTIVE
To investigate fusion expression between Bacillus thuringiensis crystal protein and a foreign protein, the expression of a fusion protein comprised of Cry1Ac, and enhanced green fluorescent protein (EGFP) in B. thuringiensis Cry(-)B strain was examined.


RESULTS
The N-terminal fusion expression of EGFP in Cry1Ac was attempted under the control of the native cry1Ac promoter. The EGFP gene was cloned into pProMu and named pProMu-EGFP. The transformant, ProMu-EGFP/CB produced parasporal inclusions that were of bipyramidal-shaped crystals in size ranging from 200 to 300 nm. The fusion protein was approximately 150 kDa and identified by the immunoblot analysis using a Cry1Ac antibody and also a GFP antibody. The LC(50) of the ProMu-EGFP/CB was twofold higher when compared with that by the ProAc/CB. However, the crystal protein produced by the ProMu-EGFP/CB was effective on Plutella xylostella larvae.


CONCLUSIONS
The ProMu-EGFP/CB produced bipyramidal shaped and insecticidal crystals comprising fusion proteins.


CONCLUSIONS
Through the N-terminal fusion expression of EGFP and Cry1Ac, expression and crystallization between the B. thuringiensis crystal protein and a foreign protein were validated.
Roh
J Y
JY
School of Agricultural Biotechnology, Seoul National University, Seoul, Korea.
Li
M S
MS
Chang
J H
JH
Choi
J Y
JY
Shim
H J
HJ
Shin
S C
SC
Boo
K S
KS
Je
Y H
YH
D000818:Animals; D001413:Bacillus thuringiensis / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001427:Bacterial Toxins; D004731:Endotoxins / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D049452:Green Fluorescent Proteins; D006460:Hemolysin Proteins; D002479:Inclusion Bodies / Q000378:metabolism / Q000648:ultrastructure; D007814:Larva / Q000254:growth & development / Q000382:microbiology; D007915:Lepidoptera / Q000254:growth & development / Q000382:microbiology*; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism*; D010572:Pest Control, Biological; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004731:Endotoxins; D006460:Hemolysin Proteins; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D049452:Green Fluorescent Proteins