Mutant pore

Patent metadata

Field	Value
Publication number	US-11939359-B2
Application number	US-202117384889-A
Country	US
Kind code	B2
Filing date	Jul 26, 2021
Priority date	Apr 6, 2016
Publication date	Mar 26, 2024
Grant date	Mar 26, 2024

Abstract

The invention relates to mutant forms of lysenin. The invention also relates to analyte characterisation using the mutant forms of lysenin.

First claim

The invention claimed is:

1. An apparatus comprising a transmembrane protein pore inserted into an in vitro membrane, wherein the transmembrane protein pore comprises at least one mutant lysenin monomer comprising a variant of the sequence shown set forth in SEQ ID NO: 2, wherein the variant comprises the K45T/R amino acid substitution.

2. The apparatus of claim 1 , wherein the transmembrane protein pore is a homo-oligomeric pore, and wherein each monomer of the homo-oligomeric pore is the mutant lysenin monomer.

3. The apparatus of claim 1 , wherein the apparatus comprises a hetero-oligomeric pore, and wherein at least one monomer of the hetero-oligomeric pore is the mutant lysenin monomer.

4. An apparatus for characterizing a target analyte, the apparatus comprising a plurality of mutant lysenin monomers, wherein each mutant lysenin monomer comprises a variant of the sequence set forth in SEQ ID NO: 2, wherein the variant comprises the K45T/R amino acid substitution, and wherein the plurality of mutant lysenin monomers is inserted into a plurality of in vitro membranes.

5. The apparatus of claim 4 , wherein each of the plurality of in vitro membranes comprises a homo-oligomeric pore consisting of the mutant lysenin monomers.

6. The apparatus of claim 4 , wherein each of the plurality of in vitro membranes comprises a hetero-oligomeric pore, and wherein at least one monomer of the hetero-oligomeric pore is the mutant lysenin monomer.

7. A transmembrane protein pore comprising at least one munt lysenin monomer, wherein the lysenin monomer comprises the amino acid sequence of SEQ ID NO: 2 having the K45T/R amino acid substitution.

8. The pore of claim 7 , wherein the pore is a hetero-oligomeric pore.

9. The pore of claim 7 , wherein the pore is a homo-oligomeric pore.

10. An apparatus produced by a method comprising: (i) obtaining a transmembrane protein pore comprising at least one mutant lysenin monomer, wherein the lysenin monomer comprises the amino acid sequence of SEQ ID NO: 2 having the K45T/R amino acid substitution; and (ii) contacting the pore with an in vitro membrane such that the pore is inserted into the in vitro membrane.

11. The apparatus of claim 1 , wherein the variant further comprises one or more of the following substitutions:
E94 D/Q/G/A/K/R; or
S49 L/K.

12. The apparatus of claim 4 , wherein each variant further comprises one or more of the following substitutions:
E94 D/Q/G/A/K/R; or
S49 UK.

13. The transmembrane protein pore of claim 7 , wherein the mutant lysenin monomer further comprises one or more of the following substitutions: E94 D/Q/G/A/K/R; or S49 UK.

14. The apparatus of claim 10 , wherein the lysenin monomer further comprises one or more of the following substitutions: E94 D/Q/G/A/K/R; or S49 UK.

Assignees

• Oxford Nanopore Tech Plc

Inventors

• Jayasinghe Lakmal
• Bruce Mark
• Mcneill Luke
• Nathani Ramiz Iqbal
• Singh Pratik Raj
• Wood Neil Roger
• Young Stephen Robert

Classifications

• C07K14/43504

  from invertebrates · CPC title

• C12Q1/6869 Primary

  Methods for sequencing · CPC title

• C07K14/43536 Primary

  from worms · CPC title

• A61K35/62

  Leeches; Worms, e.g. cestodes, tapeworms, nematodes, roundworms, earth worms, ascarids, filarias, hookworms, trichinella or taenia · CPC title

• G01N27/26

  by investigating electrochemical variables; by using electrolysis or electrophoresis · CPC title