Molecule sensor systems
The present described inventions relate, inter alia, to methods and compositions that provide for improved detection of target molecules.
1. A method for detecting a target molecule that binds to an allosteric DNA-binding protein sensor and/or switch, comprising:
(a) providing a host cell with a nucleic acid encoding the allosteric DNA-binding protein sensor and/or switch, the allosteric DNA-binding protein sensor and/or switch comprising an amino acid sequence of about 97% identity to TetR (SEQ ID NO: 46) is capable of binding apigenin, and a reporter gene system;
(b) selecting for a cell comprising the TetR sequence and the reporter gene system; and
(c) interrogating cells comprising the TetR sequence and the reporter gene system for reporter response,
the reporter response being indicative of target molecule binding to the allosteric DNA-binding protein sensor and/or switch.
2. The method of claim 1 , wherein the allosteric DNA-binding protein sensor and/or switch has one or more mutations S135A, G1381, H139M, and E147W.
3. The method of claim 1 , wherein the allosteric DNA-binding protein sensor and/or switch has one or more mutations S135H, G138A, H139M, T141A, E147Y, and H151L.
4. The method of claim 1 , wherein the allosteric DNA-binding protein sensor and/or switch has one or more mutations L131M, Y132R, S135A, G138H, H139L, E147Y, D148A, and H151L.
5. The method of claim 2 , wherein the allosteric DNA-binding protein sensor and/or switch comprises an amino acid sequence having about 97% identity to one or more of SEQ ID NOs: 48, 50, and 51.
6. A method for detecting a target molecule that binds to an allosteric DNA-binding protein sensor and/or switch, comprising:
(a) providing a host cell with a nucleic acid encoding the allosteric DNA-binding protein sensor and/or switch, the allosteric DNA-binding protein sensor and/or switch comprising an amino acid sequence of about 97% identity to TetR (SEQ ID NO: 46) is capable of binding resveratrol and has the mutation R104Y, and a reporter gene system;
(b) selecting for a cell comprising the TetR sequence and the reporter gene system; and
(c) interrogating cells comprising the TetR sequence and the reporter gene system for reporter response,
the reporter response being indicative of target molecule binding to the allosteric DNA-binding protein sensor and/or switch.
7. A method for detecting a target molecule that binds to an allosteric DNA-binding protein sensor and/or switch, comprising:
(a) providing a host cell with a nucleic acid encoding the allosteric DNA-binding protein sensor and/or switch, the allosteric DNA-binding protein sensor and/or switch comprising an amino acid sequence of about 97% identity to TetR (SEQ ID NO: 46) is capable of binding humulene and has one or more mutations G124V, G138W, E147H, and H151L, and a reporter gene system;
(b) selecting for a cell comprising the TetR sequence and the reporter gene system; and
(c) interrogating cells comprising the TetR sequence and the reporter gene system for reporter response,
the reporter response being indicative of target molecule binding to the allosteric DNA-binding protein sensor and/or switch.
8. A method for detecting a target molecule that binds to an allosteric DNA-binding protein sensor and/or switch, comprising:
(a) providing a host cell with a nucleic acid encoding the allosteric DNA-binding protein sensor and/or switch, the allosteric DNA-binding protein sensor and/or switch comprising an amino acid sequence of about 97% identity to TetR (SEQ ID NO: 46) is capable of binding nootkatone, and a reporter gene system;
(b) selecting for a cell comprising the TetR sequence and the reporter gene system; and
(c) interrogating cells comprising the TetR sequence and the reporter gene system for reporter response,
the reporter response being indicative of target molecule binding to the allosteric DNA-binding protein sensor and/or switch.
9. The method of claim 8 , wherein the allosteric DNA-binding protein sensor and/or switch comprises an amino acid sequence having about 97% identity to one or more of SEQ ID NOs: 61, 62, 63, and 64.