Biosynthesis of amorpha-4,11-diene
AssigneeThe Regents of the University of California
InventorsJay Keasling, Vincent Martin, Douglas Pitera, Sydnor T. Withers III, Jack Newman
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Abstract
Methods for synthesizing amorpha-4,11-diene synthase from isopentenyl pyrophosphate are provided. A first method comprises introducing into a host microorganism a plurality of heterologous nucleic acid sequences, each coding for a different enzyme in the mevalonate pathway for producing isopentenyl pyrophosphate. Amorpha-4,11-diene synthase is then produced using an optimized amorpha-4,11-diene synthase gene. The invention also provides nucleic acid sequences, enzymes, expression vectors, and transformed host cells for carrying out the methods.
Claims37
A method for synthesizing amorpha-4,11-diene in a host microorganism comprising:
- (a) introducing into the host microorganism a plurality of heterologous nucleic acid sequences, each coding for a different enzyme in the mevalonate pathway for producing isopentenyl pyrophosphate;
- (b) introducing into the host microorganism a DNA fragment coding for an enzyme capable of converting isopentenyl pyrophosphate to dimethylallyl pyrophosphate;
- (c) introducing into the host microorganism a DNA fragment coding for a farnesyl pyrophosphate synthase gene;
- and
- (d) introducing into the host microorganism a DNA fragment coding for an optimized synthetic amorpha-4,11-diene synthase gene.
A method for synthesizing amorpha-4,11-diene in a host microorganism, comprising:
- (a) introducing into the host microorganism an intermediate in the mevalonate pathway and at least one heterologous nucleic acid sequence, each said sequence coding for an enzyme in the mevalonate pathway necessary for converting the intermediate into isopentenyl pyrophosphate;
- (b) introducing into the host microorganism a DNA fragment coding for an enzyme capable of converting isopentenyl pyrophosphate to dimethylallyl pyrophosphate;
- (c) introducing into the host microorganism a DNA fragment coding for a farnesyl pyrophosphate synthase gene;
- and
- (d) introducing into the host microorganism a DNA fragment coding for an optimized synthetic amorpha-4,11-diene synthase gene.
A DNA fragment encoding a synthetic amorpha-4,11-diene synthase gene set forth in SEQ ID NO:37.
A DNA fragment encoding a synthetic epi-cedrol synthase gene set forth in SEQ ID NO:39.
Dependencies are inferred from the claim text.