Methods of producing polyketide synthase mutants and compositions and uses thereof
AssigneeThe Salk Institute for Biological Studies
InventorsJoseph P. Noel, Michael B Austin, Marianne E Bowman
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Abstract
The present invention comprises crystalline polyketide synthases, isolated non-native polyketide synthases having the structural coordinates of said crystalline polyketide synthases, and nucleic acids encoding such non-native polyketide synthases. Also disclosed are methods of producing mutant polyketide synthases, and methods of altering the activity and/or substrate specificity of putative polyketide synthases.
Claims66
A method of producing a mutant polyketide synthase comprising:
- (a) comparing a crystal structure of a wild type polyketide synthase with a crystal structure of a second polyketide synthase;
- (b) substituting one or more amino acids of the wild type polyketide synthase with the amino acid residues at homologous positions in the second polyketide synthase;
- and
- (c) producing said mutant polyketide synthase.
A method of producing a mutant polyketide synthase, said method comprising:
- expressing a mutant polyketide synthase created by substituting one or more amino acids of a wild type polyketide synthase with the amino acid residues at homologous positions of a second polyketide synthase, wherein said amino acid residues are selected by comparing a crystal structure of the wild type polyketide synthase with a crystal structure of the second polyketide synthase.
A method of producing a mutant polyketide synthase, said method comprising:
- synthesizing a mutant polyketide synthase created by substituting one or more amino acids of a wild type polyketide synthase with the amino acid residues at homologous positions of a second polyketide synthase, wherein said amino acid residues are selected by comparing a crystal structure of the wild type polyketide synthase with a crystal structure of the second polyketide synthase.
An isolated polyketide synthase comprising SEQ ID NO:1, wherein one or more amino acid residues are modified at one or more positions selected from the group consisting of positions 96, 98, 99, 100, 131, 133, 134, 135, 137, 157, 158, 159, 160, 165, 255, 257, 258, 266, 268, 269, 270 and 273.
A method of altering the substrate specificity of a polyketide synthase comprising:
- (a) comparing a crystal structure of a wild type polyketide synthase with a crystal structure of a second polyketide synthase;
- and
- (b) substituting one or more amino acids in the active site of the wild type polyketide synthase with the amino acid residues at homologous positions in the second polyketide synthase.
A method of altering the activity of a polyketide synthase comprising:
- (a) comparing a crystal structure of a wild type polyketide synthase with a crystal structure of a second polyketide synthase;
- and
- (b) substituting one or more amino acids of the wild type polyketide synthase with the amino acid residues at homologous positions in the second polyketide synthase.
A computer program on a computer readable medium, said computer program comprising instructions to cause a computer to:
- (a) define two different polyketide synthases or fragments thereof based on two sets of atomic coordinates derived from crystals of said two different polyketide synthases;
- and
- (b) compare the structure of said two different polyketide synthases.
Dependencies are inferred from the claim text.