Methods of developing terpene synthase variants
AssigneeAmyris, Inc.
InventorsLishan Zhao, Lan Xu, Patrick Westfall, Andrew Main
Life of this patent
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Abstract
The present disclosure relates to methods of developing terpene synthase variants through engineered host cells. Particularly, the disclosure provides methods of developing terpene synthase variants with improved in vivo performance that are useful in the commercial production of terpene products. Further encompassed in the present disclosure are superior terpene synthase variants and host cells comprising such terpene synthase variants.
Claims28
-30. (canceled)
A method of testing a terpene synthase variant for improved in vivo performance, comprising the steps of:
- (a) dividing a population of host cells into a control population and a test population;
- (b) expressing in the control population a heterologous control terpene synthase that converts a polyprenyl diphosphate to a terpene, wherein the growth rate of the host cells is dependent on having an amount of intracellular polyprenyl diphosphate;
- (c) expressing in the test population a heterologous test terpene synthase, wherein the test terpene synthase is a variant of the control terpene synthase that can convert the same polyprenyl diphosphate to the same terpene as the control terpene synthase;
- and
- (d) determining the growth rate of the control population and the test population,
- wherein the test terpene synthase is identified as having improved in vivo performance compared to the control terpene synthase by a decreased growth rate of the test population compared to the control population.
A composition comprising two cell subpopulations derived from a common population of host cells, wherein:
- (a) the first subpopulation comprises a heterologous control terpene synthase that converts a polyprenyl diphosphate to a terpene, wherein the growth rate of the host cells is dependent on having an amount of intracellular polyprenyl diphosphate;
- and
- (b) the second subpopulation comprises a heterologous test terpene synthase that is a variant of the control terpene synthase that can convert the same polyprenyl diphosphate to the same terpene as the control terpene synthase.
Dependencies are inferred from the claim text.