Methods of polypeptide identification, and compositions therefor

Patent metadata

Field	Value
Publication number	US-9528984-B2
Application number	US-201314100518-A
Country	US
Kind code	B2
Filing date	Dec 9, 2013
Priority date	Feb 13, 2006
Publication date	Dec 27, 2016
Grant date	Dec 27, 2016

Abstract

Methods are disclosed for identifying one or more proteins or polypeptides comprised by a sample. The methods comprise determining binding of each polypeptide with respect to each binding pool of a plurality of binding pools, wherein each binding pool comprises one or more probes which bind a structure comprised by a protein or polypeptide. In some aspects, polypeptides can be denatured and separated into individual polypeptide strands and immobilized on a solid support prior to determining binding of the binding pools. A protein, polypeptide or polypeptide strand can be identified by searching, in at least one database, for a protein or polypeptide sequence comprising binding pool targets either identical to or most similar to the binding pool targets comprised by the protein, polypeptide or polypeptide strand to be identified. Kits for identifying proteins, polypeptides and polypeptide strands are also disclosed.

First claim

What is claimed is:

1. A method of identifying a protein or polypeptide strand in a sample, the method comprising:
immobilizing, on a solid support, a plurality of protein or polypeptide strands comprised by a sample, such that the protein or polypeptide strand to be identified is separated by an optically resolvable distance from other protein or polypeptide strands comprised by the sample;
determining presence or absence, in the protein or polypeptide strand to be identified, of each binding pool target of a plurality of binding pool targets, wherein each binding pool target comprises one or more binding targets for one or more probes comprised by a binding pool wherein each binding pool comprises one or more probes, and binds a binding pool target comprised by a population of proteins or polypeptides described in a database;
contacting the protein or polypeptide strand to be identified sequentially or simultaneously with each binding pool;
for each binding pool, detecting binding of at least one probe comprised by the binding pool to at least one binding target if the at least one binding target is comprised by the protein or polypeptide strand to be identified;
assigning a digital signature to the protein or polypeptide strand to be identified which represents the presence or absence of binding pool targets comprised by the protein or polypeptide strand to be identified; and
identifying, in at least one database, a protein or polypeptide having a digital signature with the shortest Hamming distance to that of the protein or polypeptide strand to be identified.

2. The method of claim 1 , wherein the protein or polypeptide of the at least one database having a digital signature with the shortest Hamming distance to that of the protein or polypeptide strand to be identified comprises binding pool targets identical to those of the protein or polypeptide strand to be identified.

3. The method of claim 1 , wherein a binding target of a probe comprised by a binding pool comprises a structure selected from the group consisting of a peptide motif comprising at least one amino acid up to about six amino acids, a glycosyl moiety of a glycopeptide or glycoprotein, a GPI anchor, a disulfide linkage, a pyroglutamic acid, a nitrotyrosine and a combination thereof.

4. The method of claim 1 , wherein the plurality of binding pools comprises at least about 10 binding pools.

5. The method of claim 1 , wherein the at least one database is a protein database comprising, at least about 90% of the polypeptide sequences expressed by a species.

6. The method of claim 5 , wherein the at least one database comprises polypeptide sequences of a mammalian proteome.

7. The method of claim 1 , wherein each probe is selected from the group consisting of at least one antibody, at least one aptamer, at least one kinase, at least one avimer and any combination thereof.

8. The method of claim 1 , wherein detecting binding of at least one probe comprised by the binding pool comprises detecting a label bound directly or indirectly to the at least one probe.

9. The method of claim 1 , wherein the protein or polypeptide having a digital signature that has the shortest Hamming distance to that of the protein or polypeptide strand to be identified comprises identifying a sequence comprised by at least one database having a digital signature that is identical to the digital signature of the protein or polypeptide strand to be identified.

10. The method of claim 1 , further comprising, denaturing the protein or polypeptide strand to be identified.

11. The method of claim 1 , wherein the solid support comprises an optically transparent medium selected from the group consisting of glass and non-fluorescent plastic.

12. The method of claim 1 , wherein the immobilizing the polypeptide strands comprises (i) covalently coupling the protein or polypeptide strands to the solid support and/or (ii) contacting the protein or polypeptide strands with a surface of the solid support, and coating the surface with a porous gel.

13. The method of claim 1 , further comprising quantifying the protein or polypeptide strands comprising the same or substantially the same (i) binding pool targets or (ii) digital signatures, wherein a digital signature of a protein or polypeptide strand is derived from the presence or absence of binding, pool targets in the protein or polypeptide strand.

14. The method of claim 1 , wherein the optically resolvable distance is about 0.5 micron or greater.

15. The method of claim 11 , wherein the solid support comprises a reactive moiety which covalently binds the protein or polypeptide strands.

Assignees

• Univ Washington
• Mitra Robi D

Inventors

• Mitra Robi D

Classifications

• G01N33/54306 Primary

  Solid-phase reaction mechanisms · CPC title

• G01N33/582

  with fluorescent label · CPC title

• G01N33/6845 Primary

  Methods of identifying protein-protein interactions in protein mixtures · CPC title