Mutant thioesterases
Mutant thioesterases having enhanced medium chain substrate activity, polynucleotides encoding and configured to express the mutant thioesterases in a transformed host cell, host cells transformed to contain the polynucleotides, and methods of using same.
1. An unnatural, mutated protein comprising an amino acid sequence at least about 80% identical to positions 19-317 of SEQ ID NO:4, wherein:
the amino acid sequence comprises:
a residue other than asparagine at a position corresponding to position 28 of SEQ ID NO:4; and
a residue other than isoleucine at a position corresponding to position 65 of SEQ ID NO:4;
the protein lacks an N-terminal portion having an amino acid sequence identical to positions 1-18 of SEQ ID NO:4; and
the protein exhibits thioesterase activity.
2. The protein of claim 1 , wherein the amino acid sequence at least about 80% identical to positions 19-317 of SEQ ID NO:4 comprises:
a serine, cysteine, threonine, tyrosine, or glutamine at the position corresponding to position 28 of SEQ ID NO:4; and
a methionine, alanine, leucine, phenylalanine, valine, proline, or glycine at the position corresponding to position 65 of SEQ ID NO:4.
3. The protein of claim 2 , wherein the protein comprises, relative to a protein consisting of an amino acid sequence of SEQ ID NO:4, an N-terminal truncation of positions 1-18 of SEQ ID NO:4.
4. The protein of claim 1 , wherein the amino acid sequence at least about 80% identical to positions 19-317 of SEQ ID NO:4 comprises:
a serine at the position corresponding to position 28 of SEQ ID NO:4; and
a methionine at the position corresponding to position 65 of SEQ ID NO:4.
5. The protein of claim 4 , wherein the protein comprises, relative to a protein consisting of an amino acid sequence of SEQ ID NO:4, an N-terminal truncation of positions 1-18 of SEQ ID NO:4.
6. The protein of claim 1 , wherein the protein comprises, relative to a protein consisting of an amino acid sequence of SEQ ID NO:4, an N-terminal truncation of positions 1-18 of SEQ ID NO:4.
7. The protein of claim 1 , wherein the amino acid sequence at least about 80% identical to positions 19-317 of SEQ ID NO:4 is an amino acid sequence at least about 85% identical to positions 19-317 of SEQ ID NO:4.
8. The protein of claim 7 , wherein the amino acid sequence at least about 85% identical to positions 19-317 of SEQ ID NO:4 comprises:
a serine, cysteine, threonine, tyrosine, or glutamine at the position corresponding to position 28 of SEQ ID NO:4; and
a methionine, alanine, leucine, phenylalanine, valine, proline, or glycine at the position corresponding to position 65 of SEQ ID NO:4.
9. The protein of claim 8 , wherein the protein comprises, relative to a protein consisting of an amino acid sequence of SEQ ID NO:4, an N-terminal truncation of positions 1-18 of SEQ ID NO:4.
10. The protein of claim 7 , wherein the amino acid sequence at least about 85% identical to positions 19-317 of SEQ ID NO:4 comprises:
a serine at the position corresponding to position 28 of SEQ ID NO:4; and
a methionine at the position corresponding to position 65 of SEQ ID NO:4.
11. The protein of claim 10 , wherein the protein comprises, relative to a protein consisting of an amino acid sequence of SEQ ID NO:4, an N-terminal truncation of positions 1-18 of SEQ ID NO:4.
12. The protein of claim 7 , wherein the protein comprises, relative to a protein consisting of an amino acid sequence of SEQ ID NO:4, an N-terminal truncation of positions 1-18 of SEQ ID NO:4.
13. The protein of claim 1 , wherein the amino acid sequence at least about 80% identical to positions 19-317 of SEQ ID NO:4 is an amino acid sequence at least about 90% identical to positions 19-317 of SEQ ID NO:4.
14. The protein of claim 13 , wherein the amino acid sequence at least about 90% identical to positions 19-317 of SEQ ID NO:4 comprises:
a serine, cysteine, threonine, tyrosine, or glutamine at the position corresponding to position 28 of SEQ ID NO:4; and
a methionine, alanine, leucine, phenylalanine, valine, proline, or glycine at the position corresponding to position 65 of SEQ ID NO:4.
15. The protein of claim 14 , wherein the protein comprises, relative to a protein consisting of an amino acid sequence of SEQ ID NO:4, an N-terminal truncation of positions 1-18 of SEQ ID NO:4.
16. The protein of claim 13 , wherein the amino acid sequence at least about 90% identical to positions 19-317 of SEQ ID NO:4 comprises:
a serine at the position corresponding to position 28 of SEQ ID NO:4; and
a methionine at the position corresponding to position 65 of SEQ ID NO:4.
17. The protein of claim 16 , wherein the protein comprises, relative to a protein consisting of an amino acid sequence of SEQ ID NO:4, an N-terminal truncation of positions 1-18 of SEQ ID NO:4.
18. The protein of claim 13 , wherein the protein comprises, relative to a protein consisting of an amino acid sequence of SEQ ID NO:4, an N-terminal truncation of positions 1-18 of SEQ ID NO:4.
19. The protein of claim 1 , wherein the amino acid sequence at least about 80% identical to positions 19-317 of SEQ ID NO:4 is an amino acid sequence at least about 95% identical to positions 19-317 of SEQ ID NO:4.
20. The protein of claim 19 , wherein the amino acid sequence at least about 95% identical to positions 19-317 of SEQ ID NO:4 comprises:
a serine, cysteine, threonine, tyrosine, or glutamine at the position corresponding to position 28 of SEQ ID NO:4; and
a methionine, alanine, leucine, phenylalanine, valine, proline, or glycine at the position corresponding to position 65 of SEQ ID NO:4.
21. The protein of claim 20 , wherein the protein comprises, relative to a protein consisting of an amino acid sequence of SEQ ID NO:4, an N-terminal truncation of positions 1-18 of SEQ ID NO:4.
22. The protein of claim 19 , wherein the amino acid sequence at least about 95% identical to positions 19-317 of SEQ ID NO:4 comprises:
a serine at the position corresponding to position 28 of SEQ ID NO:4; and
a methionine at the position corresponding to position 65 of SEQ ID NO:4.
23. The protein of claim 22 , wherein the protein comprises, relative to a protein consisting of an amino acid sequence of SEQ ID NO:4, an N-terminal truncation of positions 1-18 of SEQ ID NO:4.
24. The protein of claim 19 , wherein the protein comprises, relative to a protein consisting of an amino acid sequence of SEQ ID NO:4, an N-terminal truncation of positions 1-18 of SEQ ID NO:4.