Fluorescent leptin sensor protein

Abstract

One object of this disclosure is to provide a protein whose fluorescence increases when bound to leptin. [Solution] Starting from the N-terminus, (a) the N-terminal fragment of the fluorescent protein, (b) the first linker, (c) the C-terminal fragment of the leptin-binding domain derived from the leptin receptor, (d) the synthetic linker, The present invention provides a protein comprising (e) the N-terminal fragment of a leptin-binding domain derived from a leptin receptor, (f) a second linker, and (g) the amino acid sequence of the C-terminal fragment of a fluorescent protein, wherein when the C-terminal fragment of the leptin-binding domain derived from a leptin receptor and the N-terminal fragment of the leptin-binding domain derived from a leptin receptor bind to leptin, the fluorescence emitted from the N-terminal fragment of the fluorescent protein and the C-terminal fragment of the fluorescent protein increases.

Claims

1. Starting from the N-terminus, (a) N-terminal fragment of fluorescent protein, (b) First linker, (c) C-terminal fragment of leptin-binding domain derived from leptin receptor, (d) Synthetic linker, (e) N-terminal fragment of leptin-binding domain derived from leptin receptor, (f) Second linker, and (g) C-terminal fragment of a fluorescent protein A protein containing the following amino acid sequence, in which the C-terminal fragment of the leptin receptor-derived leptin-binding domain and the N-terminal fragment of the leptin receptor-derived leptin-binding domain bind to leptin, thereby increasing the fluorescence emitted from the N-terminal fragment and the C-terminal fragment of the fluorescent protein.

2. The C-terminal fragment of the leptin-binding domain derived from the leptin receptor consists of an amino acid sequence that has approximately 90% or more identity with the amino acid sequence of any of sequence numbers 15, 17, and 18. The N-terminal fragment of the leptin receptor-derived leptin-binding domain consists of an amino acid sequence that has approximately 90% or more identity with the amino acid sequence of SEQ ID NO: 16 or 19. The protein according to claim 1.

3. The protein according to claim 1, wherein the fluorescent protein is EGFP or a variant thereof.

4. The N-terminal fragment of the fluorescent protein consists of an amino acid sequence that has approximately 90% or more identity with the amino acid sequence of SEQ ID NO: 8 or 9. The C-terminal fragment of the fluorescent protein consists of an amino acid sequence that is approximately 90% or more identical to the amino acid sequence of SEQ ID NO:

10. The protein according to claim 1.

5. The protein according to claim 1, wherein the synthetic linker consists of an amino acid sequence having approximately 90% or more identity with the amino acid sequence of SEQ ID NO: 20 or 21.

6. The protein according to claim 1, comprising a cell membrane localization leader sequence at its N-terminus.

7. The protein according to claim 1, comprising a cell membrane anchor sequence at its C-terminus.

8. A protein containing any of the amino acid sequences from sequence numbers 30 to 32.

9. Starting from the N-terminus, (a) C-terminal fragment of leptin-binding domain derived from leptin receptor, (b) Synthetic linker, (c) N-terminal fragment of leptin-binding domain derived from leptin receptor, A leptin-binding protein containing the following amino acid sequence.

10. The C-terminal fragment of the leptin-binding domain derived from the leptin receptor consists of an amino acid sequence that has approximately 90% or more identity with the amino acid sequence of any of sequence numbers 15, 17, and 18. The N-terminal fragment of the leptin receptor-derived leptin-binding domain consists of an amino acid sequence that has approximately 90% or more identity with the amino acid sequence of SEQ ID NO: 16 or 19. The leptin-binding protein according to claim 9.

11. The leptin-binding protein according to claim 9, wherein the synthetic linker consists of an amino acid sequence having approximately 90% or more identity with the amino acid sequence of SEQ ID NO: 20 or 21.

12. A leptin-binding protein containing one of the amino acid sequences of sequence numbers 38-40.

13. A nucleic acid encoding a protein according to any one of claims 1 to 12.

14. A vector comprising the nucleic acid described in claim 13.

15. A cell containing the nucleic acid described in claim 13.

16. A method for detecting leptin in a sample, Contacting the protein described in any of claims 1 to 8 with the sample, To evaluate the fluorescence emitted from the aforementioned protein, Methods that include...

17. A method for measuring leptin in a sample, Contacting the protein described in any of claims 1 to 8 with the sample, To measure the fluorescence intensity of the fluorescence emitted from the aforementioned protein, Methods that include...

18. A kit for detecting or measuring leptin, comprising the protein described in any one of claims 1 to 8.

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