Methods and device for analyte characterization

• US 8,278,055 B2
• Filed: 10/29/2003
• Issued: 10/02/2012
• Est. Priority Date: 05/01/2002
• Status: Active Grant

• Current Assignee: Intel Corporation
• Original Assignee: Intel Corporation
• Inventors: Su, Xing, Berlin, Andrew A.
• Primary Examiner(s): Ha, Julie
• Application Number:
  US10/697,682
• Publication Number:
  US 20050282229A1
• Time in Patent Office: 3,261 Days
• Field of Search: None
• US Class Current: 435/7.1
• CPC Class Codes: G01N 33/48721 Investigating individual macromolecules, e.g. by translocation through nanopores Coulter counters in general G01N15/12; fabrication methods for nanoscale apertures B81B1/00; sequencing of nucleic acids C12Q1/68
• CPC Class Codes: G01N 33/582 with fluorescent label
• CPC Class Codes: G01N 33/6803 General methods of protein analysis not limited to specific proteins or families of proteins
• CPC Class Codes: G01N 33/6818 Sequencing of polypeptides

Abstract

The methods and apparatus, disclosed herein are of use for sequencing and/or identifying proteins, polypeptides and/or peptides. Proteins containing labeled amino acid residues may be synthesized and passed through nanopores. A detector operably coupled to a nanopore may detect labeled amino acid residues as they pass through the nanopore. Distance maps for each type of labeled amino acid residue may be compiled. The distance maps may be used to sequence and/or identify the protein. Apparatus of use for protein sequencing and/or identification is also disclosed herein. In alternative methods, other types of analytes may be analyzed by the same techniques.

Claims

1. A method comprising:

a) placing a plurality of labeled proteins, polypeptides or peptides in a plurality of chambers, such that different chambers contain a different type of labeled amino acid;

b) passing the labeled proteins, polypeptides or peptides through one or more nanopores, an inner surface of the nanopores coated with a semiconductor material;

c) detecting labeled amino acid residues in the labeled proteins, polypeptides or peptides;

d) compiling an amino acid distance map for each type of labeled amino acid; and

e) identifying the protein based on the distance maps.

2. The method of claim 1, further comprising:

a) placing a template nucleic acid into each chamber; and

b) producing the one or more labeled proteins, polypeptides or peptides encoded by the template nucleic acid.

3. The method of claim 1, further comprising:

a) obtaining one or more proteins, polypeptides or peptides from a biological sample; and

b) labeling the proteins, polypeptides or peptides post-translationally.

4. The method of claim 1, wherein the protein, polypeptide or peptide is identified by comparing the distance maps with a library of amino acid distance maps.

5. The method of claim 1, wherein the protein, polypeptide or peptide is identified by comparing the distance maps with the sequences of known proteins.

6. The method of claim 2, wherein each chamber is operably coupled to a different set of nanopores.

7. The method of claim 1, wherein each nanopore is operably coupled to a detector.

8. The method of claim 1, wherein only one labeled protein, polypeptide or peptide passes through a nanopore at a time.

9. The method of claim 1, wherein the length of time between passage of a first labeled amino acid through the nanopore and passage of a second labeled amino acid through the nanopore corresponds to the distance along the labeled protein, polypeptide or peptide between the first and second amino acids.

10. The method of claim 1, wherein the labels are selected from the group consisting of luminescent labels, fluorescent labels, phosphorescent labels, chemiluminescent labels, conductive labels, nuclear magnetic resonance labels, mass spectroscopy labels, electron spin resonance labels, electron paramagnetic resonance labels and Raman labels.

11. The method of claim 1, wherein at least one end of the labeled protein, polypeptide or peptide is attached to an identifiable label.

12. The method of claim 1, wherein said labeled amino acids are detected with a photodetector.

13. The method of claim 1, wherein said labeled amino acids are detected with an electrical detector.

14. The method of claim 2, further comprising analyzing a multiplicity of labeled proteins, polypeptides or peptides from each chamber.

15. The method of claim 1, further comprising determining at least a partial sequence of the protein, polypeptide or peptide based on the distance maps.

16. The method of claim 2, wherein the one or more labeled proteins, polypeptides or peptides encoded by the template nucleic acid is produced by in vitro translation or by linked transcription/translation.

17. The method of claim 16, wherein in vitro translation is performed with mRNA templates.

18. The method of claim 16, wherein in vitro translation carried out in cell systems of rabbit reticulocyte lysates, wheat germ extracts, or E. coli extracts.

19. The method of claim 1, wherein the distance map shows distances in a sub-nanometer scale.