Chemical modification of glucose oxidase and its application to biosensors

• US 10,190,100 B1
• Filed: 12/28/2015
• Issued: 01/29/2019
• Est. Priority Date: 12/28/2015
• Status: Active Grant

• Current Assignee: Verily Life Sciences LLC (Alphabet Inc.)
• Original Assignee: Verily Life Sciences LLC (Alphabet Inc.)
• Inventors: Liu, Zenghe, Zhou, Cindy Xiaoxin
• Primary Examiner(s): Desai, Anand U
• Application Number:
  US14/981,723
• Time in Patent Office: 1,128 Days
• Field of Search: None
• CPC Class Codes: C12N 9/0006 acting on CH-OH groups as donors (1.1)
• CPC Class Codes: C12Q 1/006 for glucose
• CPC Class Codes: C12Y 101/03004 Glucose oxidase (1.1.3.4)
• CPC Class Codes: G01N 2333/904 acting on CHOH groups as donors, e.g. glucose oxidase, lactate dehydrogenase (1.1)
• CPC Class Codes: G01N 33/54353 with ligand attached to the carrier via a chemical coupling agent coatings G01N33/54393

Abstract

A modified glucose oxidase for use in a sensor for the continuous or semi-continuous monitoring of glucose is disclosed. The modified glucose oxidase may include a glucose oxidase having at least one amino group substituted with a methacrylate through a hydrophilic linker including at least one alkylene oxide unit. A glucose sensor is also disclosed. The glucose sensor includes a crosslinked, hydrophilic copolymer sensing layer in contact with a surface of an electrode, where the sensing layer includes methacrylate-derived backbone chains covalently bound to glucose oxidase through a hydrophilic linker including at least one alkylene oxide unit. Also included is a method for making the modified glucose oxidase and the glucose sensor.

Claims

1. A modified glucose oxidase comprising a glucose oxidase wherein at least one amino group is substituted with a methacrylate through a hydrophilic linker comprising at least one ethylene or propylene oxide unit.

2. The modified glucose oxidase of claim 1, wherein the linker comprises at least one ethylene oxide unit.

3. The modified glucose oxidase of claim 1, having the structure of formula (Ia):

4. The modified glucose oxidase of claim 1, having the structure of formula (Ib):

5. The modified glucose oxidase of claim 4, wherein the M_n of the poly(ethylene oxide) portion of the modified glucose falls within the range of 300 to 400.

6. The modified glucose oxidase of claim 1, having the structure of formula (Ic):

7. The modified glucose oxidase of claim 6, wherein n is 10-20;

m is 30-50; and

l is 1-50.

8. The modified glucose oxidase of claim 6, wherein the combined M_n of the poly(ethylene oxide) and poly(propylene oxide) portion of the modified glucose falls within the range of about 4,000 g/mol to about 5,000 g/mol.

9. A method for making the modified glucose oxidase of claim 1, the method comprising reacting glucose oxidase with an activated methacrylate, wherein the methacrylate is bound through a linker comprising at least one ethylene or propylene oxide unit to a reactive group capable of forming a covalent bound to glucose oxidase.

10. A glucose sensor comprising:

a sensing layer in contact with a surface of an electrode, wherein the sensing layer comprises;

backbone chains comprising

first methacrylate-derived monomeric units, each of which is covalently bound to a primary amino group on glucose oxidase through a hydrophilic linker comprising at least one ethylene or propylene oxide unit,

second methacrylate-derived monomeric units, and

optionally, third methacrylate-derived monomeric units, each having a hydrophilic side chain,

hydrophilic crosslinks between the second methacrylate-derived units in different backbone chains; and

optionally an additional layer provided on the sensing layer comprising third methacrylate-derived monomeric units, each having a hydrophilic side chain,

wherein the third methacrylate-derived monomeric units are present in at least one of the additional layer or the backbone chains of the sensing layer.

11. The sensor of claim 10, wherein the third methacrylate-derived monomeric units are present in the backbone chains of the sensing layer.

12. The sensor of claim 10, wherein the third methacrylate-derived monomeric units are present in an additional layer, wherein the additional layer is a protective membrane comprising:

backbone chains comprising;

third methacrylate-derived monomeric units, each unit having a hydrophilic side chain; and

fourth methacrylate-derived monomeric units; and

hydrophilic crosslinks between the fourth methacrylate-derived units in different backbone chains.

13. The sensor of claim 10, wherein the first methacrylate-derived monomeric units have the structure of formula (IIa):

14. The sensor of claim 10, wherein the first methacrylate-derived monomeric units have the structure of formula (IIb):

15. The sensor of claim 10, wherein the first methacrylate-derived monomeric units have the structure of formula (IIc):

16. The sensor of claim 10, wherein the third methacrylate-derived units have the structure of formula (IIIa):

17. The sensor of claim 10, wherein the third methacrylate-derived units have the structure:

18. The sensor of claim 12, wherein the hydrophilic crosslinks between the second or fourth methacrylate-derived units in different backbone chains are derived from the di(ethylene oxide) portion of di(ethylene glycol) dimethacrylate.

19. The sensor of claim 10, wherein the sensing layer comprises:

first methacrylate-derived monomeric units having the structure of formula (IIb);

20. The sensor of claim 10, wherein the sensing layer comprises:

first methacrylate-derived monomeric units having the structure of formula (IIc);