Part:BBa_K5143016
A Bt toxin secreted in Saccharomyces cerevisiae that binds to cellulose
Description
Plu1537 is a 14kDa insecticidal toxin protein derived from Photorhabdus luminescens TT01. It exhibits strong toxicity against termites through oral ingestion. With 30% homology to a known Bt toxin, it is believed to function in a similar manner, likely by disrupting the gut cells of the insects. However, our experiments have shown that this protein has not been effective in termite control when administered directly, suggesting that the mode of delivery is critical for its efficacy.
In our project, we are developing a versatile insect trap using the Plu1537 toxin. This toxin is expressed in yeast as part of a co-culture system, where it is fused with a Cellulose-Binding Domain (CBD) and an alpha factor for secretion. The CBD allows the toxin to attach to cellulose produced by a bacterium also present in the co-culture. When termites consume the cellulose, they ingest the toxin as well. This method ensures that the toxin is effectively delivered to the termites through the cellulose, making the trap highly effective and targeted for termite control. Additionally, by fusing the Plu1537 toxin with a CBD and an alpha factor, we can enhance its stability and attachment to the cellulose matrix, ensuring a higher ingestion rate by the termites.
Details of the System Function
The schematic illustrates the design of our insect trap system. The Plu1537 toxin is genetically fused to an alpha factor, which facilitates the secretion of the protein from yeast cells, and a Cellulose-Binding Domain (CBD), which enables the toxin to bind firmly to cellulose. The yeast cells expressing this chimeric protein are cultured together with cellulose-producing bacteria. As the yeast secretes the chimeric protein, the CBD component binds to the cellulose produced by the bacteria, effectively functionalizing the cellulose with the toxin. When termites consume the cellulose, they also ingest the toxin, leading to their demise. This schematic highlights the innovative integration of genetic engineering and microbiological techniques to deliver the insecticidal protein in a highly targeted and efficient manner.
Construction
The gene encoding the chimeric Plu1537 protein fused to alpha-factor and CBD has been optimized for synthesis and expression in Saccharomyces cerevisiae. We have placed this coding region downstream of the GAP promoter to ensure strong constitutive expression. The entire construct has been synthesized.
This genetic construct has been cloned into the following plasmid backbone: BBa_K514005
Resulting in the following integrative plasmid:
References
iGEM ZJU-China 2015
M. Li et al., MOL BIOL REP 36, 785 (2009).
M. S. Kelker et al., PLOS ONE 9, (2014).
BBa_K1321340
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23INCOMPATIBLE WITH RFC[23]Unknown
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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