Claims
- 1. A method for selecting an enzyme mutant with desired catalytic activity, said method comprising:
displaying each of a multitude of mutants of an enzyme on a surface of a display vehicle, saiddisplay vehicle comprising a nucleic acid selecting for a display vehicle carrying a nucleic acid encoding an enzyme mutant having at least a mutated site other than the catalytic site and having the desired activity.
- 2. The method according to claim 1, wherein said mutated site comprises a substrate binding site or allosteric site of said enzyme.
- 3. The method according to claim 1 or 2, further comprising selecting for improved binding of said enzyme displayed on said surface with a selected substrate for said enzyme.
- 4. The method according to claim 1 or 2, further comprising selecting for improved binding of said enzyme displayed on said surface with an effector for said enzyme.
- 5. The method according to claim 3 or 4, wherein said binding occurs under a set environmental condition.
- 6. The method according to claim 5, wherein said environmental condition comprises a condition capable of influencing binding and/or said catalytic activity.
- 7. The method according to claim 6, wherein said condition capable of influencing binding and/or said catalytic activity is selected from the group comprising the presence or concentration of an electrolyte, salt, substrate or product, heterologous enzyme or effector, or temperature.
- 8. The method according to claim 5 or 6, wherein said set environmental condition comprises a set pH.
- 9. The method according any one of the above claims, wherein said binding of said enzyme displayed on said surface is tested in at least two rounds of selection.
- 10. The method according to claim 9, wherein said display vehicle is not amplified between at least two rounds of selection.
- 11. A mutant enzyme selectable according to any one of claims 1 to 10.
- 12. The mutant enzyme of claim 11 having a catalytic site that is at least partly distinct from its substrate binding site or its allosteric site.
- 13. The mutant enzyme of claim 11 or claim 12, further comprising a polysaccharide binding enzyme.
- 14. The mutant enzyme of any one of claims 11 to 13, having improved catalytic activity over the wild-type enzyme's catalytic activity at a set environmental condition.
- 15. The mutant enzyme of claim 14, wherein said environmental condition is the pH value under which catalytic activity is determined.
- 16. The mutant enzyme of any one of claims 11 to 15, comprising an α-amylase.
- 17. The mutant enzyme of claim 16, derived from Bacillus licheniformis.
- 18. The mutant enzyme of claim 16 or claim 17, further comprising improved catalytic activity over its wild-type enzyme at pH 4.5.
- 19. A nucleic acid encoding the mutant enzyme of any one of claims 11 to 18.
- 20. A vector comprising the nucleic acid of claim 19.
- 21. A host cell comprising the vector of claim 20.
- 22. A host cell comprising the nucleic acid according to claim 19.
- 23. A process to obtain a desired product, said process comprising altering a substrate by subjecting said substrate to catalytic activity of an enzyme of any one of claims 11 to 18.
- 24. The process of claim 23, wherein said substrate comprises a polysaccharide.
- 25. A product produced by the process of claim 23 or claim 24.
- 26. A process for selecting a mutant protein or mutant part thereof, said process comprising:
displaying each of a multitude of mutants of a protein or part thereof on a surface of a display vehicle, said display vehicle comprising a nucleic acid encoding a mutant of said protein or part thereof; and selecting for a display vehicle carrying nucleic acid encoding a mutant of said protein or part thereof by successive rounds of selection wherein said display vehicle is not amplified between at least two successive rounds of selection.
- 27. A mutant protein or mutant part thereof selected by the process of claim 26.
- 28. A nucleic acid encoding the protein or part thereof of claim 27.
- 29. A vector comprising the nucleic acid of claim 28.
- 30. A host cell comprising the vector of claim 29.
- 31. A host cell comprising the nucleic acid of claim 28.
Priority Claims (1)
Number |
Date |
Country |
Kind |
99201846.5 |
Jun 1999 |
EP |
|
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a continuation of co-pending International Patent Application No. PCT/NL00/00399 filed on Jun. 9, 2000 designating the United States of America (International Publication No. WO 00/77193 published on Dec. 21, 2000), the contents of the entirety of which are incorporated by this reference, which claims the benefit under 35 U.S.C. §119(e) of U.S. Provisional Application No. 60/138,443, filed Jun. 10, 1999.
Provisional Applications (1)
|
Number |
Date |
Country |
|
60138443 |
Jun 1999 |
US |
Continuations (1)
|
Number |
Date |
Country |
Parent |
PCT/NL00/00399 |
Jun 2000 |
US |
Child |
10015979 |
Dec 2001 |
US |