Described are methods for the production of isobutene comprising the enzymatic conversion of 3-methylcrotonic acid into isobutene wherein the enzymatic conversion of 3-methylcrotonic acid into isobutene is achieved by making use of an FMN-dependent decarboxylase associated with an FMN prenyl transferase, wherein said FMN prenyl transferase catalyzes the prenylation of a flavin cofactor (FMN or FAD) utilizing dimethylallyl phosphate (DMAP) into a flavin-derived cofactor, wherein said method further comprises providing said DMAP enzymatically by: (i) the enzymatic conversion of dimethylallyl pyrophosphate (DMAPP) into said DMAP; or (ii) a single enzymatic step in which prenol is directly enzymatically converted into said DMAP; or (iii) two enzymatic steps comprising: first enzymatically converting DMAPP into prenol; and then enzymatically converting the thus obtained prenol into said DMAP; or (iv) the enzymatic conversion of isopentenyl monophosphate (IMP) into said DMAP, or by a combination of any one of (i) to (iv). Moreover, described are methods for the production of isobutene comprising the enzymatic conversion of 3-methylcrotonic acid into isobutene wherein the enzymatic conversion of 3-methylcrotonic acid into isobutene is achieved by making use of an FMN-dependent decarboxylase associated with an FMN prenyl transferase, wherein said FMN prenyl transferase catalyzes the prenylation of a flavin cofactor (FMN or FAD) utilizing dimethylallyl pyrophosphate (DMAPP), wherein said method further comprises providing said DMAPP enzymatically by: (v) the enzymatic conversion of isopentenyl pyrophosphate (IPP) into said DMAPP; or (vi) the enzymatic conversion of dimethylallyl phosphate (DMAP) into said DMAPP; or (vii) the enzymatic conversion of prenol into said DMAPP; (viii) or by a combination of any one of (v) to (vii). Moreover, described are methods for providing said flavin cofactor enzymatically by the enzymatic conversion of riboflavin into flavin mononucleotide (FMN).

The present invention concerns a method of producing and enantiomerically pure alpha-ionone. Further, the invention concerns a nucleic acid that comprises a sequence that encodes a lycopene-epsilon-cyclase (EC), a lycopene-epsilon-cyclase (EC), plasmids, which encode components of the alpha-ionone biosynthesis and a microorganism that contains heterologous nucleotide sequences which encode the enzymes geranylgeranyl-diphosphate-synthase, isopentenyl-diphosphate-isomerase (IPI), phytoene desaturase-dehydrogenase (crtI), phytoene synthase (crtB), lycopene-epsilon-cyclase (EC) and carotenoid-cleavage-dioxygenase (CCD1). Further, the invention concerns a method of producing highly pure epsilon-carotene.

The present disclosure relates to the use of a maltose dependent degron to control stability of a protein of interest fused thereto at the post-translational level. The present disclosure also relates to the use of a maltose dependent degron in combination with a maltose-responsive promoter to control gene expression at the transcriptional level and to control protein stability at the post-translational level. The present disclosure also relates to the use of a stabilization construct that couples expression of a cell-growth-affecting protein with the production of non-catabolic compounds. The present disclosure further relates to the use of a synthetic maltose-responsive promoter. The present disclosure further provides compositions and methods for using a maltose dependent degron, a maltose-responsive promoter, and a stabilization construct, either alone or in various combinations, for the production of non-catabolic compounds in genetically modified host cells.

The present disclosure relates to the use of a maltose dependent degron to control stability of a protein of interest fused thereto at the post-translational level. The present disclosure also relates to the use of a maltose dependent degron in combination with a maltose-responsive promoter to control gene expression at the transcriptional level and to control protein stability at the post-translational level. The present disclosure also relates to the use of a stabilization construct that couples expression of a cell-growth-affecting protein with the production of non-catabolic compounds. The present disclosure further relates to the use of a synthetic maltose-responsive promoter. The present disclosure further provides compositions and methods for using a maltose dependent degron, a maltose-responsive promoter, and a stabilization construct, either alone or in various combinations, for the production of non-catabolic compounds in genetically modified host cells.

The invention provides a method for producing a terpene or a precursor thereof by microbial fermentation. Typically, the method involves culturing a recombinant bacterium in the presence of a gaseous substrate whereby the bacterium produces a terpene or a precursor thereof, such as mevalonic acid, isopentenyl pyrophosphate, dimethylallyl pyrophosphate, isoprene, geranyl pyrophosphate, farnesyl pyrophosphate, and/or farnesene. The bacterium may comprise one or more exogenous enzymes, such as enzymes in mevalonate, DXS, or terpene biosynthesis pathways.

Cells comprising a heterologous metabolic pathway are configured to produce a terpene product containing non-multiples of five carbon, particularly wherein the pathway comprises heterologous Lepidoptera insect juvenile hormone biosynthetic pathway enzymes of the insect's mevalonate pathway.

The invention relates to microbial terpene production. Known methods for microbial production of terpenes are mostly based on the direct conversion of sugars. Therefore alternative substrates, in particular alternative carbon sources, for use in microbial terpene production were desirable. The invention relates to a methylotrophic bacterium containing recombinant DNA coding for at least one polypeptide with enzymatic activity for heterologous expression in said bacterium, wherein said at least one polypeptide with enzymatic activity is selected from the group consisting an enzyme of a heterologous mevalonate pathway, a heterologous terpene synthase and optionally a heterologous synthase of a prenyl diphosphate precursor. The invention further relates in particular to a method for de novo microbial synthesis of sesquiterpenes or diterpenes from methanol and/or ethanol.

Methods and compositions for synthesizing dienes and derivative thereof, such as isoprene, in Cupriavidus necator are provided.

The invention features methods for producing isoprene from cultured cells. The invention also provides compositions that include these cultured cells.

The invention provides a method for producing an isoprenoid or a precursor thereof by microbial fermentation. Typically, the method involves culturing a recombinant bacterium in the presence of a gaseous substrate whereby the bacterium produces an isoprenoid or a precursor thereof, such as mevalonic acid, isopentenyl pyrophosphate, dimethylallyl pyrophosphate, isoprene, geranyl pyrophosphate, farnesyl pyrophosphate, and/or pinene. The bacterium may comprise one or more exogenous enzymes, such as enzymes in mevalonate, DXS, isoprenoid alcohol, or terpene biosynthesis pathways.