Binding protein as biosensors

• US 6,855,556 B2
• Filed: 01/04/2002
• Issued: 02/15/2005
• Est. Priority Date: 01/04/2002
• Status: Expired due to Fees

• Current Assignee: Becton, Dickinson & Co
• Original Assignee: Becton, Dickinson & Co
• Inventors: Wright, David J., Sherman, Douglas B., Pitner, J. Bruce, Amiss, Terry J., Nycz, Colleen M.
• Primary Examiner(s): ALEXANDER, LYLE
• Application Number:
  US10/040,077
• Publication Number:
  US 20030134346A1
• Time in Patent Office: 1,138 Days
• Field of Search: 436/56, 436/86, 436/95, 436/164, 436/172, 435/14, 435/287.1, 435/7.2, 435/817, 435/814
• US Class Current: 436/95
• CPC Class Codes: C07K 14/245 Escherichia (G)
• CPC Class Codes: G01N 33/5438 Electrodes
• CPC Class Codes: Y10S 435/817 Enzyme or microbe electrode
• CPC Class Codes: Y10T 436/144444 Glucose

Abstract

The invention is directed to compositions of mutated binding proteins containing reporter groups, analyte biosensor devices derived there from, and their use as analyte biosensor both in vitro and in vivo.

Claims

1. A glucose biosensor comprising at least one mutated glucose/galactose binding protein and at least one reporter group attached to said binding protein, such that said reporter group provides a detectable and reversible signal change when said mutated binding protein is exposed to varying glucose concentrations;

wherein said at least one mutated glucose/galactose binding protein comprises at least one amino acid substitution selected from the group consisting of a cysteine at position 11, a cysteine at position 14, a cysteine at position 19, a cysteine at position 43, a cysteine at position 74, a cysteine at position 107, a cysteine at position 110, a cysteine at position 112, a cysteine at position 113, a cysteine at position 137, a cysteine at position 149, a cysteine at position 213, a cysteine at position 216, a cysteine at position 238, a cysteine at position 287, and a cysteine at position 292, and

wherein said detectable and reversible signal change is related to said varying concentrations.

2. The biosensor of claim 1 wherein said reporter group is a luminescent label.

3. The biosensor of claim 1, wherein said mutated glucose/galactose binding protein has at least one histidine tag.

4. The biosensor of claim 2 wherein said luminescent label is covalently coupled to said at least one mutated glucose/galactose binding protein.

5. The biosensor of claim 4 wherein said luminescent label has an excitation wavelength of more than about 600 nanometers.

6. The biosensor of claim 4 wherein said luminescent label has an emission wavelength of more than about 600 nanometers.

7. The biosensor of claim 4, wherein said luminescent label is selected from the group consisting of fluorescein, coumarins, rhodamines, 5-TMRIA (tetramethylrhodamine-5-iodoacetamide), (9-(2(or 4)-(N-(2-maleimdylethyl)-sulfonamidyl)-4(or 2)-sulfophenyl)-2,3,6,7,12,13,16,17-octahydro-(1H,5H,11H,15-xantheno(2,3,4-ij:

5,6,7-i′

j′

)diquinolizin-18-ium salt), 2-(5-(1-(6-(N-(2-maleimdylethyl)-amino)-6-oxohexyl)-1,3-dihydro-3,3-dimethyl-5-sulfo-2H-indol-2-ylidene )-1,3-propyldienyl)-1-ethyl-3,3-dimethyl-5-sulfo-3H-indolium salt, N-((2-iodoacetoxy)ethyl)-N-methyl)am-ino-7-nitrobenzoxadiazole, 6-acryloyl-2-dimethylaminonaphthalene, pyrene, 6-amino-2,3-dihydro-2-(2-((iodoacetyl)amino)ethyl)-1,3-dioxo-1H-benz(de)isoquinoline-5,8-disulfonic acid salt, 2-(5-(1-(6-N-(2-maleimdylethyl)-amino)-6-oxobexyl)-1,3-dihydro-3,3-dimethyl-5-sulfo-2H-indol-2ylidene)-1,3-pentadienyl)-1-ethyl-3,3-dimethyl-5-sulfo-3H-indolium salt, 4-(5-(4-dimethylaminophenyl)oxazole-2-yl)-N-(2-bromoacetamidoethyl)sulfonamide, (N-(4,4-difluoro-1,3,5,7-tetramethyl-4-bora-3a,4a-diaza-s-indacene-2-yl)iodoacetamide, N-(4,4-difluoro-5,7-diphenyl-4-bora-3a,4a-diaza-s-indacene-3-propionyl)-N-′

-iodoacetylethylenediarnine, 5-((((2-iodoacetyl)amino)ethyl)amino)naphthalene-1-sulfonic acid, and carboxy-X-rhodamine, 5/6-iodoacetamide.

8. A glucose biosensor comprising at least one mutated glucose/galactose binding protein and at least one reporter group attached to said binding protein, such that said reporter group provides a detectable and reversible signal change when said mutated binding protein is exposed to varying glucose concentrations;

wherein said at least one mutated glucose/galactose binding protein comprises at least two amino acid substitutions, said at least two amino acid substitutions being selected from the group consisting of a cysteine at position 112 and a serine at position 238, a cysteine at position 149 and a serine at position 238, a cysteine at position 152 and a cysteine at position 182, a cysteine at position 152 and a serine at position 213, a cysteine at position 213 and a cysteine at position 238, a cysteine at position 149 and an arginine at position 213; and

wherein said detectable and reversible signal change is related to said varying concentrations.

9. The biosensor of claim 8 wherein said reporter group is a luminescent label.

10. The biosensor of claim 8, wherein said mutated glucose/galactose binding protein has at least one histidine tag.

11. The biosensor of claim 9 wherein said luminescent label is covalently coupled to said at least one mutated glucose/galactose binding protein.

12. The biosensor of claim 11 wherein said luminescent label has an excitation wavelength of more than about 600 nanometers.

13. The biosensor of claim 11 wherein said luminescent label has an emission wavelength of more than about 600 nanometers.

14. The biosensor of claim 11, wherein said luminescent label is selected from the group consisting of fluorescein, coumarins, rhodamines, 5-TMRIA (tetramethylrhodamine-5-iodoacetamide), (9-(2(or 4)-(N-(2-maleimdylethyl)-sulfonamidyl)-4(or 2)-sulfophenyl)-2,3,6,7,12,13,16,17-octahydro-(1H,5H,11H,15H-xantheno(2,3,4-ij:

5,6,7-i′

j′

)diquinolizin-18-ium salt), 2-(5-(1-(6-(N-(2-maleimdylethyl)-amino)-6-oxohexyl)-1,3-dihydro-3,3-dimethyl-5-sulfo-2H-indol-2-ylidene )-1,3-propyldienyl)-1-ethyl-3,3-dimethyl-5-sulfo-3H-indolium salt, N-((2-iodoacetoxy)ethyl)-N-methyl)am-ino-7-nitrobenzoxadiazole, 6-acryloyl-2-dimethylaminonaphthalene, pyrene, 6-amino-2,3-dihydro-2-(2-((iodoacetyl)amino)ethyl)-1,3-dioxo-1H-benz(de)isoquinoline-5,8-disulfonic acid salt, 2-(5-(1-(6-N-(2-maleimdylethyl)-amino)-6-oxohexyl)-1,3-dihydro-3,3-dimethyl-5-sulfo-2H-indol-2-ylidene)-1,3-pentadienyl)-1-ethyl-3,3-dimethyl-5-sulfo-3H-indolium salt, 4-(5-(4dimethylaminophenyl)oxazole-2-yl)-N-(2-bromoacetamidoethyl)sulfonamide, (N-(4,4-difluoro-1,3,5,7-tetramethyl-4-bora-3a,4a-diaza-s-indacene-2-yl)iodoacetamide, N-(4,4-difluoro-5,7-diphenyl-4-bora-3a,4a-diaza-s-indacene-3-propionyl)-N-′

-iodoacetylethylenediamine, 5-((((2-iodoacetyl)amino)ethyl)amino)naphthalene-1-sulfonic acid, and carboxy-X-rhodamine, 5/6-iodoacetamide.

15. A glucose biosensor comprising at least one mutated glucose/galactose binding protein and at least one reporter group attached to said binding protein, such that said reporter group provides a detectable and reversible signal change when said mutated binding protein is exposed to varying glucose concentrations;

wherein said at least one mutated glucose/galactose binding protein comprises at least three amino acid substitutions, said at least three amino acid substitutions being selected from the group consisting of a cysteine at position 149 and a serine at position 213 and a serine at position 238, and a cysteine at position 149 and an arginine at position 213 and a serine at position 238; and

wherein said detectable and reversible signal change is related to said varying concentrations.

16. The biosensor of claim 15 wherein said reporter group is a luminescent label.

17. The biosensor of claim 15 wherein said mutated glucose/galactose binding protein has at least one histidine tag.

18. The biosensor of claim 16 wherein said luminescent label is covalently coupled to said at least one mutated glucose/galactose binding protein.

19. The biosensor of claim 18 wherein said luminescent label has an excitation wavelength of more than about 600 nanometers.

20. The biosensor of claim 18 wherein said luminescent label has an emission wavelength of more than about 600 nanometers.

21. The biosensor of claim 18, wherein said luminescent label is selected from the group consisting of fluorescein, coumarins, rhodamines, 5-TMRIA (tetramethylrhodamine-5-iodoacetamide), (9-(2(or 4)-(N-(2-maleimdylethyl)-sulfonamidyl)-4(or 2)-sulfophenyl)-2,3,6,7,12,13,16,17-octahydro-(1H,5H,11H,15H-xantheno(2,3,4-ij:

5,6,7-i′

j′

)diquinolizin-18-ium salt), 2-(5-(1-(6-(N-(2-maleimdylethyl)-amino)-6-oxohexyl)-1,3-dihydro-3,3-dimethyl-5-sulfo-2H-indol-2-ylidene)-1,3-propyldienyl)-1-ethyl-3,3-dimethyl-5-sulfo-3H-indolium salt, N-((2-iodoacetoxy)ethyl)-N-methyl)am-ino-7-nitrobenzoxadiazole, 6-acryloyl-2-dimethylaminonaphthalene, pyrene, 6-amino-2,3-dihydro-2-(2-((iodoacetyl)amino)ethyl)-1,3-dioxo-1H-benz(de)isoquinoline-5,8-disulfonic acid salt, 2-(5-(1-(6-N-(2-maleimdylethyl)-amino)-6-oxohexyl)-1,3-dihydro-3,3-dimethyl-5-sulfo-2H-indol-2-ylidene)-1,3-pentadienyl)-1-ethyl-3,3-dimethyl-5-sulfo-3H-indolium salt, 4-(5-(4-dimethylaminophenyl)oxazole-2-yl)-N-(2-bromoacetamidoethyl)sulfonamide, (N-(4,4-difluoro-1,3,5,7-tetramethyl-4-bora-3a,4a-diaza-s-indacene-2-yl)iodoacetamide N-(4,4-difluoro-1,3,5,7-diphenyl-4-bora-3a, 4a -diaza-s-indacene-3-propionyl)-N-′

-iodoacetylethylenediamine, 5-((((2-iodoacetyl)amino)ethyl)amino)naphthalene 1-sulfonic acid, and carboxy-X-rhodamine, 5/6-iodoacetamide.