Gene construct encoding mutant Thioesterase, mutant Thioesterase encoded thereby, transformed host cell containing the gene construct, and method of using them to produce medium-chain fatty acids
AssigneesWisconsin Alumni Research Foundation, The Penn State Research Foundation
InventorsBrian F. Pfleger, Nestor Jose Hernandez-Lozada, Costas Maranas, Matthew Grisewood
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Abstract
Unnatural, mutated thioesterases having an amino acid sequence that is at least 80% identical to SEQ. ID. NO: 1 and having substitutions at one or more of amino acid positions I107, R108, L109, S122, M141, E142, Y145, and L146, gene constructs encoding and configured to express the mutated thioesterases in a transformed host cell and host cells transformed to contain the gene constructs.
Claims15
An unnatural, mutated protein comprising an amino acid sequence that is at least 85% identical to SEQ ID NO:1 and comprises one or more substitutions with respect to SEQ ID NO:1, wherein the protein has at least one of enhanced thioesterase activity and enhanced thioesterase specificity in catalyzing the hydrolysis of a medium-chain acyl-acyl carrier protein substrate or a medium-chain acyl-CoA substrate to yield a free fatty acid or a free fatty acid derivative compared to an unaltered protein of SEQ ID NO:1, wherein the one or more substitutions comprise one or more of:
- a lysine at a position aligning to R108 of SEQ ID NO:1;
- a tyrosine at a position aligning to F139 of SEQ ID NO:1;
- a valine at a position aligning to 142 of SEQ ID NO:1;
- a glycine at a position aligning to A111 of SEQ ID NO:1;
- a serine at a position aligning to P138 of SEQ ID NO:1;
- an isoleucine at a position aligning to M170 of SEQ ID NO:1;
- a serine at a position aligning to T35 of SEQ ID NO:1;
- and
- an aspartate as a position aligning to E142 of SEQ ID NO:1.
Dependencies are inferred from the claim text.