Uranium-chelating peptides derived from EF-hand calcium-binding motif useful for uranium biodetection and biodecontamination

Patent metadata

Field	Value
Publication number	US-9845343-B2
Application number	US-201414779114-A
Country	US
Kind code	B2
Filing date	Mar 28, 2014
Priority date	Mar 28, 2013
Publication date	Dec 19, 2017
Grant date	Dec 19, 2017

Abstract

Uranium-chelating polypeptides comprising at least one helix-loop-helix calcium-binding (EF-hand) motif which comprises a deletion of at least two amino acid in the 12-amino-acid calcium-binding loop sequence, and their use for uranium biodetection and biodecontamination.

First claim

The invention claimed is:

1. A polypeptide comprising at least one helix-loop-helix calcium-binding (EF-hand) motif with deletion of two amino acid residues in the 12-amino-acid calcium-binding loop sequence selected from the group consisting of deletion in positions 1 and 2, and deletion in positions 2 and 3, wherein said polypeptide binds uranyl.

2. The polypeptide of claim 1 , wherein said EF-hand motif(s) are derived from signaling EF-hand protein(s) of the calmodulin superfamily selected from the group consisting of calmodulin and troponin C.

3. The polypeptide of claim 1 , which comprises two or four EF-hand motifs, wherein at least one EF-hand motif comprises said deletion in the calcium-binding loop sequence, and the other EF-hand motif(s) comprise said deletion or not.

4. The polypeptide of claim 2 , which is a calmodulin domain 1 variant comprising two EF-hand motifs, respectively from the EF-hand 1 and the EF-hand2 of calmodulin protein(s).

5. The polypeptide of claim 3 , which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 17, 18, 20 and 60.

6. A fusion protein comprising a polypeptide according to claim 1 fused to another protein moiety.

7. The fusion protein of claim 6 , which is a cameleon protein comprising tandem fusions of a fluorescence-donor protein, a polypeptide which comprises two or four EF-hand motifs, wherein at least one EF-hand motif comprises said deletion in the calcium-binding loop sequence and the other EF-hand motif(s) comprise said deletion or not, and a fluorescence-acceptor protein.

8. The fusion protein of claim 7 , which comprises a sequence selected from the group consisting of SEQ ID NO: 35, 38, 61, 63 and 67.

9. The polypeptide of claim 1 or a fusion protein comprising a polypeptide according to claim 1 fused to another protein moiety, which is immobilized onto a solid support.

10. A polynucleotide encoding the polypeptide of claim 1 .

11. A host cell comprising with the polynucleotide of claim 10 , wherein the host cell is not an organism.

12. A non-human transgenic organism comprising the polynucleotide of claim 10 .

13. A method of detecting uranium contamination in a sample comprising contacting the sample with a uranyl chelating agent comprising:
i. the polypeptide of claim 1 ,
ii. a fusion protein comprising a polypeptide according to claim 1 fused to another protein moiety,
iii. a host cell comprising a polynucleotide encoding the polypeptide of claim 1 or a fusion protein comprising a polypeptide according to claim 1 fused to another protein moiety, or
iv. A non-human transgenic organism comprising a polynucleotide encoding the polypeptide of claim 1 or a fusion protein comprising a polypeptide according to claim 1 fused to another protein moiety, and
detecting the presence of uranyl chelation in the sample.

14. A method of decontaminating or bio-remediating a sample containing uranium comprising contacting the sample with a uranyl chelating agent comprising:
i. the polypeptide of claim 1 ,
ii. a fusion protein comprising a polypeptide according to claim 1 fused to another protein moiety,
iii. a host cell comprising a polynucleotide encoding the polypeptide of claim 1 or a fusion protein comprising a polypeptide according to claim 1 fused to another protein moiety, or
iv. A non-human transgenic organism comprising a polynucleotide encoding the polypeptide of claim 1 or a fusion protein comprising a polypeptide according to claim 1 fused to another protein moiety.

15. The polypeptide of claim 3 , which consists of an amino acid sequence selected from the group consisting of SEQ ID NO: 17, 18, 20 and 60.

16. The fusion protein of claim 7 , which consists of a sequence selected from the group consisting of SEQ ID NO: 35, 38, 61, 63 and 67.

17. A polynucleotide encoding a fusion protein comprising the polypeptide according to claim 1 fused to another protein moiety.

Assignees

• Commissariat Energie Atomique

Inventors

• Pardoux Romain
• Sauge-Merle Sandrine
• Lemaire David
• Berthomieu Catherine
• Guilbaud Philippe
• Delangle Pascale
• Bremond Nicolas
• Beccia Maria-Rosa

Classifications

• C07K14/4728 Primary

  Calcium binding proteins, e.g. calmodulin · CPC title

• G21F9/307

  in polymeric matrix, e.g. resins, tars · CPC title

• C07K2319/60

  containing spectroscopic/fluorescent detection, e.g. green fluorescent protein [GFP] · CPC title

• G01N33/20

  Metals · CPC title