Immobilized transaminases and process for making and using immobilized transaminase
The invention is directed to immobilized transaminases and methods of making and using them.
1. A resin comprising an immobilized transaminase polypeptide, wherein the transaminase polypeptide comprises the amino acid sequence of SEQ ID NO: 1, wherein the resin comprises methacrylate, and wherein the transaminase polypeptide is attached to the resin.
2. The resin of claim 1 , wherein the transaminase is attached to the resin by hydrophilic interactions.
3. The resin of claim 1 , wherein the resin comprises DIAION HP2MG.
4. The resin of claim 3 , wherein the transaminase is stable in a solvent system that comprises at least 90% organic solvent.
5. The resin of claim 3 , wherein the transaminase is stable in a solvent system that comprises at least 95% organic solvent.
6. The resin of claim 2 , wherein the resin comprises methacrylate with no functional groups.
7. The resin of claim 2 , wherein the resin comprises methacrylate with no aromatic groups.
8. A resin comprising an immobilized transaminase polypeptide, wherein the transaminase polypeptide comprises the amino acid sequence of SEQ ID NO: 1, wherein the resin comprises DIAION HP2MG, and wherein the transaminase polypeptide is physically attached to the resin via hydrophilic interactions.
9. A process for making (2R)-4-oxo-4-[3-(trifluoromethyl)-5,6-dihydro[1,2,4]triazolo[4,3-a]pyrazin-7(8H)-yl]-1-(2,4,5-trifluorophenyl)butan-2-amine comprising the steps of:
1) dissolving 4-oxo-4-[3-(trifluoromethyl)-5,6-dihydro[1,2,4]triazolo[4,3-a]pyrazin-7(8H)-yl]-1-(2,4,5-trifluorophenyl)butan-2-one in an organic solvent; and
2) contacting the 4-oxo-4-[3-(trifluoromethyl)-5,6-dihydro[1,2,4]triazolo[4,3-a]pyrazin-7(8-H)-yl]-1-(2,4,5-trifluorophenyl)butan-2-one with the resin comprising the immobilized transaminase of claim 1 and an amino group donor to thereby make (2R)-4-oxo-4-[3-(trifluoromethyl)-5,6-dihydro[1,2,4]triazolo[4,3-a]pyrazi-n-7(8H)-yl]-1-(2,4,5-trifluorophenyl)butan-2-amine.
10. The process of claim 9 wherein the immobilized transaminase is physically attached to the resin, and the resin is DIAION HP2MG.
11. A process of making a resin comprising an immobilized transaminase comprising the steps of:
1) incubating a solution of a transaminase polypeptide with a resin to form a resin comprising an immobilized transaminase polypeptide, wherein the transaminase polypeptide comprises the amino acid sequence of SEQ ID NO: 1, wherein the resin comprises methacrylate, and wherein the transaminase polypeptide is attached to the resin; and
2) filtering and rinsing the immobilized transaminase.