Nucleoside triphosphate transporter and uses thereof
Disclosed herein are proteins, methods, cells, engineered microorganisms, and kits for generating a modified nucleoside triphosphate transporter from Phaeodactylum tricornutum . Also disclosed herein proteins, methods, cells, engineered microorganisms, and kits for production of a nucleic acid molecule that comprises an unnatural nucleotide utilizing a modified nucleoside triphosphate transporter from Phaeodactylum tricornutum.
1. A method of increasing production of a nucleic acid molecule in an E. coli cell, comprising:
(a) expressing in the cell a recombinant nucleoside triphosphate transporter, wherein the recombinant nucleoside triphosphate transporter has an amino acid sequence selected from SEQ ID NO: 4, SEQ ID NO: 6 and SEQ ID NO: 8,
(b) incubating the cell with one or more unnatural nucleoside triphosphates, and
(c) transporting the one or more unnatural nucleoside triphosphates into the cell using the recombinant nucleoside triphosphate transporter, wherein the nucleic acid molecule comprises one or more unnatural nucleotides comprising a base selected from the group consisting of
2. The method of claim 1 , wherein the nucleic acid molecule comprises DNA.
3. The method of claim 1 , wherein the nucleic acid molecule comprises RNA.
4. The method of claim 1 , wherein the nucleic acid molecule comprises one or more unnatural nucleotides comprising a base which is
5. The method of claim 1 , wherein the nucleic acid molecule comprises one or more unnatural nucleotides comprising a base which is
6. The method of claim 1 , wherein the one or more unnatural nucleoside triphosphates comprises a base selected from the group consisting of
7. The method of claim 1 , wherein the one or more unnatural nucleoside triphosphates comprises a base which is
8. The method of claim 1 , wherein the one or more unnatural nucleoside triphosphates comprises a base which is
9. A method of increasing production of a DNA nucleic acid molecule in an E. coli cell comprising:
(a) expressing in the cell a recombinant nucleoside triphosphate transporter, wherein the recombinant nucleoside triphosphate transporter has an amino acid sequence comprising selected from SEQ ID NO: 4, SEQ ID NO: 6 and SEQ ID NO: 8,
(b) incubating the cell with one or more unnatural 2′-deoxyribonucleoside triphosphates, and
(c) transporting the one or more unnatural 2′-deoxyribonucleoside triphosphates into the cell using the recombinant nucleoside triphosphate transporter, wherein the one or more unnatural 2′-deoxyribonucleoside triphosphates comprise a base selected from the group consisting of:
10. A method of increasing production of a RNA nucleic acid molecule in an E. coli cell, comprising:
(a) expressing in the cell a recombinant nucleoside triphosphate transporter, wherein the recombinant nucleoside triphosphate transporter has an amino acid sequence selected from SEQ ID NO: 4, SEQ ID NO: 6 and SEQ ID NO: 8,
(b) incubating the cell with one or more unnatural ribonucleoside triphosphates, and
(c) transporting the one or more unnatural ribonucleoside triphosphates into the cell using the recombinant nucleoside triphosphate transporter, wherein the one or more unnatural ribonucleoside triphosphates comprise a base selected from the group consisting of:
11. The method of claim 1 , wherein the recombinant nucleoside triphosphate transporter has amino acid sequence SEQ ID NO: 4.
12. The method of claim 11 , wherein the one or more unnatural nucleoside triphosphates comprises a base selected from
13. The method of claim 1 , wherein the recombinant nucleoside triphosphate transporter has amino acid sequence SEQ ID NO: 6.
14. The method of claim 13 , wherein the one or more unnatural nucleoside triphosphates comprises a base selected from
15. The method of claim 1 , wherein the recombinant nucleoside triphosphate transporter has amino acid sequence SEQ ID NO: 8.
16. The method of claim 15 , wherein the one or more unnatural nucleoside triphosphates comprises a base selected from
17. The method of claim 9 , wherein the recombinant nucleoside triphosphate transporter has amino acid sequence SEQ ID NO: 4.
18. The method of claim 17 , wherein the one or more unnatural 2′-deoxyribonucleoside triphosphates comprises a base selected from
19. The method of claim 9 , wherein the recombinant nucleoside triphosphate transporter has amino acid sequence SEQ ID NO: 6.
20. The method of claim 19 , wherein the one or more unnatural 2′-deoxyribonucleoside triphosphates comprises a base selected from
21. The method of claim 9 , wherein the recombinant nucleoside triphosphate transporter has amino acid sequence SEQ ID NO: 8.
22. The method of claim 21 , wherein the one or more unnatural 2′-deoxyribonucleoside triphosphates comprises a base selected from
23. The method of claim 10 , wherein the recombinant nucleoside triphosphate transporter has amino acid sequence SEQ ID NO: 4.
24. The method of claim 23 , wherein the one or more unnatural ribonucleoside triphosphates comprises a base selected from
25. The method of claim 10 , wherein the recombinant nucleoside triphosphate transporter has amino acid sequence SEQ ID NO: 6.
26. The method of claim 25 , wherein the one or more unnatural ribonucleoside triphosphates comprises a base selected from
27. The method of claim 10 , wherein the recombinant nucleoside triphosphate transporter has amino acid sequence SEQ ID NO: 8.
28. The method of claim 27 , wherein the one or more unnatural ribonucleoside triphosphates comprises a base selected from