Compositions and methods for production of myrcene
AssigneeAmyris, Inc.
InventorsAndrew MAIN, Grzegorz WOJCIECHOWSKI, Yue YANG, Lishan ZHAO
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Abstract
Provided herein are compositions and methods for producing myrcene by culturing genetically modified microbial host cells that express a myrcene synthase and optionally a geranyl pyrophosphate synthase. Also provided herein are isolated nude is acid molecules that encode myrcene synthase variants derived from the Ocimum species myrcene synthase, which comprise one or more amino acid substitutions that improve in vivo performance of myrcene synthase in genetically modified microbial host cells. Also provided herein are isolated myrcene synthase variants that exhibit an improved activity for converting geranyl diphosphate into myrcene.
Claims32
A fermentation composition comprising:
- (a) a genetically modified microbial host cell cultured in a culture medium, wherein the microbial host cell comprises a heterologous nucleic acid molecule encoding a myrcene synthase;
- and
- (b) monoterpenes produced from the microbial host cell, wherein the monoterpenes comprise myrcene as a major component and one or more co-products as minor components, wherein the one or more co-products comprise α-terpinene and γ-terpinene.
23-37. (canceled)
A genetically modified microbial host cell comprising:
- (a) a heterologous nucleic acid molecule encoding an Ocimum species myrcene synthase that comprises:
- i. an amino acid sequence of SEQ ID NO:
- 2;
- or
- ii. an amino acid sequence that has at least 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99% sequence identity to SEQ ID NO:
- 2;
- and
- (b) a heterologous nucleic acid molecule encoding a geranyl pyrophosphate synthase.
39-51. (canceled)
An isolated nucleic acid molecule encoding a myrcene synthase, wherein the myrcene synthase comprises:
- (a) an amino acid sequence that has at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% sequence identity to SEQ ID NO:
- 2;
- and
- (b) at least one variant amino acid residue compared to SEQ ID NO:
- 2 at one or more of positions selected from the group of positions consisting of positions 27, 28, 207, 213, 222, 342, 347, 381, 382, 389, 390, 401, 404, 428, 439, 466, 482, 484, 505, 514, 517, 524, 527, 528, 543, 544, and 552, wherein the positions are numbered with reference to SEQ ID NO:
- 2.
53-61. (canceled)
64-71. (canceled)
A method of producing myrcene, the method comprising:
- (a) culturing a population of a genetically modified host cell in a first culture medium under a non-inducing condition, wherein the genetically modified host cell comprises a heterologous nucleic acid molecule encoding a myrcene synthase and a heterologous nucleic acid molecule encoding a geranyl pyrophosphate synthase;
- and
- (b) culturing the population or a subpopulation thereof in a second culture medium under an inducing condition which increases production of myrcene compared to the non-inducing condition of step (a),
- wherein the second culture medium comprises monoterpenes produced from the population or subpopulation of the genetically modified host cell, wherein the monoterpenes comprise myrcene as a major component and one or more co-products as minor components, wherein the one or more co-products comprise α-terpinene and γ-terpinene.
73-76. (canceled)
Dependencies are inferred from the claim text.