Fusion of the genes for AHL-lactonase and S-layer protein in Bacillus thuringiensis increases its ability to inhibit soft rot caused by Erwinia carotovora
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Two genes, ctc and ctc2, responsible for surface layer (S-layer) protein synthesis in Bacillus thuringiensis CTC, were mutated and resulted in B. thuringiensis Tr5. To synthesize and express the N-acyl-homoserine lactonase (AHL-lactonase) in the extracellular space of B. thuringiensis, the aiiA 4Q7 gene (an AHL-lactonase gene from B. thuringiensis 4Q7), which confers the ability to inhibit plant soft rot disease in B. thuringiensis 4Q7, was fused with the upstream sequence of the ctc gene, which in turn is essential for S-layer protein secretion and anchoring on the cell surface. The resulting fusion gene, slh-aiiA, was expressed in B. thuringiensis Tr5 to avoid competition for the extracellular space with the native S-layer protein. Our results indicate that B. thuringiensis Tr5 containing the fusion gene slh-aiiA displayed high extracellular AHL-degrading activity. When compared with wild-type B. thuringiensis strains, the ability of the constructed strain to inhibit soft rot disease caused by Erwinia carotovora SCG1 was markedly increased. These findings provide evidence for a significant advance in our ability to inhibit soft rot disease caused by E. carotovora.
KeywordsN-acyl-homoserine lactonase Fusion protein AiiA4Q7 protein Bacillus thuringiensis Surface layer Soft rot disease
Strain DH5a (pJZ365) was kindly donated by Professor Stephen C. Winans in Cornell University. This study was supported by grants from the National Basic Research Program (973) of China (2003CB114201), National Natural Science Foundation of China (30270053, and 30080013), and National High Technology Research and Development project (863) of China (2006, 2004AA214092 and 2003AA223081). We thank Donald H. Dean at the Ohio State University, Chen Guo-qiang at Tsinghua University, Louis S. Tisa at University of New Hampshire, AP James Chin at the University of Queensland for their critical reading of this manuscript.
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