Chimeric receptor for use in whole-cell sensors for detecting analytes of interest
The present invention relates to chimeric receptors that can be used in whole-cell sensors for detecting analytes of interest. The inventors showed that the DNA binding domains and downstream gene expression can be activated via dimerization of an artificial dimerization composed of a single chain variable domain. They demonstrated for the first time that an artificial bacterial receptor using an antibody-like domain can be activated and produce a transcriptional output upon ligand-binding. In particular, the present invention relates to a chimeric receptor polypeptide comprising: i) a first DNA binding domain, ii) at least one binding domain selected from the group consisting of heavy chain variable domain, camelid VHHs, or antibody mimetics having specificity for an analyte, and iii) a linker between the DNA binding domain and the binding domain.
1 . A chimeric receptor polypeptide comprising (i) a first DNA binding domain, wherein the first DNA binding domain consists of an amino acid sequence with at least 75% identity with SEQ ID NO:1, SEQ ID NO: 2, SEQ ID NO: 8, or SEQ ID NO: 13, (ii) at least one binding domain having specificity for an analyte of interest, wherein the binding domain is a single domain antibody selected from the group consisting of a heavy chain variable domain and a camelid VHH, and (iii) a linker between the first DNA binding domain and the at least one binding domain, wherein the linker consists of an amino acid sequence with at least 90% identity with SEQ ID NO:3, SEQ ID NO: 4, SEQ ID NO: 5 or SEQ ID NO: 6.
2 . The chimeric receptor polypeptide of claim 1 further comprising a spacer inserted between the linker and the at least one binding domain.
3 . The chimeric receptor polypeptide of claim 2 wherein the spacer comprises the amino acid sequence DTRLPMS (SEQ ID NO:7) or GGGSG (SEQ ID NO:12).
4 . The chimeric receptor polypeptide of claim 1 , wherein the analyte of interest is a small molecule.
5 . A cell-free system comprising the chimeric receptor polypeptide of claim 1 .
6 . The cell-free system of claim 5 wherein the chimeric receptor polypeptide is embedded partially or completely in a solid support.
7 . The cell-free system of claim 6 wherein the solid support is a porous substrate.
8 . The cell-free system of claim 6 wherein the solid support is paper.
9 . The cell-free system of claim 6 wherein the solid support comprises several spatially distinct reaction regions where the cell-free system is confined.
10 . The cell-free system of claim 6 wherein the components are lyophilized on the solid support.
11 . The cell-free system of claim 5 further comprising a reporter gene.
12 . The cell-free system of claim 11 wherein the reporter gene encodes a fluorescent protein.