This document relates to using bidirectional, multi-enzymatic scaffolds to biosynthesize cannabinoids in recombinant hosts.

Described is a recombinant organism or microorganism which is capable of enzymatically converting acetyl-CoA into isobutene, (A) wherein in said organism or microorganism: (i) acetyl-CoA is enzymatically converted into acetoacetyl-CoA, (ii) acetoacetyl-CoA is enzymatically converted into 3-hydroxy-3-methylglutaryl-CoA, (iii) 3-hydroxy-3-methylglutaryl-CoA is enzymatically converted into 3-methylglutaconyl-CoA, (iv) 3-methylglutaconyl-CoA is enzymatically converted into 3-methylcrotonyl-CoA, and (v) wherein said 3-methylcrotonyl-CoA is converted into isobutene by: (a) enzymatically converting 3-methylcrotonyl-CoA into 3-methylcrotonic acid which is then further enzymatically converted into said isobutene; or (b) enzymatically converting 3-methylcrotonyl-CoA into 3-hydroxy-3-methylbutyryl-CoA which is then further enzymatically converted into 3-hydroxy-3-methylbutyric acid which is then further enzymatically converted into 3-phosphonoxy-3-methylbutyric acid which is then further enzymatically converted into said isobutene; (B) wherein said recombinant organism or microorganism has an increased pool of coenzyme A (CoA) over the organism or microorganism from which it is derived due to: (i) an increased uptake of pantothenate; and/or (ii) an increased conversion of pantothenate into CoA. Moreover, described is the use of such a recombinant organism or microorganism for the production of isobutene. Further, described is a method for the production of isobutene by culturing such a recombinant organism or microorganism in a suitable culture medium under suitable conditions.

Provided is a method of producing at least one omega-hydroxyl fatty acid, the method comprising: (a) contacting at least one alkane with at least one recombinant yeast cell in an aqueous medium, wherein the yeast cell is capable of oxidising the alkane to the corresponding omega-hydroxyl fatty acid and the yeast cell comprises a reduced fatty acid degradation capacity.

A PHA copolymer which is slowly crystallized is improved in crystallization speed to improve the melt workability of the PHA copolymer in working such as injection molding, film molding, blow molding, fiber spinning, extrusion foaming or bead foaming, thereby improving the resultant articles in productivity. A method for the improvement is a method for producing a PHA mixture, including the step of culturing a microorganism having both of a gene encoding a PHA synthase that synthesizes a copolymer PHA (A) and that is derived from the genus Aeromonas, and a gene encoding a PHA synthase that synthesizes a PHA (B) different in melting point from the copolymer PHA (A) by 10° C. or more to produce, in a cell of the microorganism, two or more PHAs different in melting point from one another by 10° C. or more simultaneously.

The invention provides non-naturally occurring microbial organisms containing a fatty alcohol, fatty aldehyde or fatty acid pathway, wherein the microbial organisms selectively produce a fatty alcohol, fatty aldehyde or fatty acid of a specified length. Also provided are non-naturally occurring microbial organisms having a fatty alcohol, fatty aldehyde or fatty acid pathway, wherein the microbial organisms further include an acetyl-CoA pathway. In some aspects, the microbial organisms of the invention have select gene disruptions or enzyme attenuations that increase production of fatty alcohols, fatty aldehydes or fatty acids. The invention additionally provides methods of using the above microbial organisms to produce a fatty alcohol, a fatty aldehyde or a fatty acid.

Provided is engineered microorganisms expressing acetoacetyl-CoA reductase variants and a method for improving the yield of PHA. Compared with the wild-type acetoacetyl-CoA reductase represented by SEQ ID NO. 31, the variant has one or more of the following mutations: (1) mutation of valine at position 141 to isoleucine or leucine; (2) mutation of methionine at position 12 to threonine, serine, alanine, leucine, lysine or isoleucine; (3) mutation of isoleucine at position 194 to valine, leucine or methionine; (4) mutation of glutamic acid at position 42 to lysine, glutamine, leucine, aspartic acid, proline, threonine, asparagine, or histidine; and (5) mutation of phenylalanine at position 55 to valine, alanine or isoleucine. The variants and their coding genes can promote the synthesis and accumulation of PHA by the strain and increase the yield of PHA.

The primary object of the present invention is to provide a polymer synthase gene which is derived from mangrove soil metagenome, and the method for producing the useful copolymer by using this polymer synthase. Another object of the present invention is to provide an enoyl-CoA hydratase gene which is derived from Streptomyces sp. CFMR 7, and the method for producing the useful copolymer, P(3HB-co-3HHx) with increasing the composition of 3HHx by the expression of this enoyl-CoA hydratase.

In order to achieve these objects, an isolated polynucleotide encoding for a polypeptide with polymer synthase activity comprising an amino acid sequence set forth in SEQ ID NO: 1 or 3, or an amino acid sequence set forth in SEQ ID NO: 1 or 3 wherein one or more amino acids is replaced, deleted or added are provided.

The invention provides an engineered carboxylic acid reductase (CAR) enzyme, a nucleic acid encoding the CAR enzyme, and a non-naturally occurring microbial organism comprising an exogenous nucleic acid encoding the CAR, an engineered transaminase (TA) enzyme, and/or a hexamethylenediamine (HMD) transaminase (TA2) enzyme. The invention provides a non-naturally occurring microbial organism that has a 1,6-hexanediol (HDO) pathway with a HDO pathway enzyme expressed in sufficient amounts to produce 6 aminocaproate semi aldehyde, HDO, or both. The invention further provides a non-naturally occurring microbial organism that has an HMD pathway with a HMD pathway enzyme expressed in sufficient amounts to produce 6-aminocaproate semialdehyde, HMD, or both. The invention additionally provides bioderived HMD, 6-aminocaproate semialdehyde, and/or HDO and methods for producing bioderived HMD, 6-aminocaproate semialdehyde, and/or HDO.

The use of microorganisms to make alpha-functionalized chemicals and fuels, (e.g. alpha-functionalized carboxylic acids, alcohols, hydrocarbons, amines, and their beta-, and omega-functionalized derivatives), by utilizing an iterative carbon chain elongation pathway that uses functionalized extender units. The core enzymes in the pathway include thiolase, dehydrogenase, dehydratase and reductase. Native or engineered thiolases catalyze the condensation of either unsubstituted or functionalized acyl-CoA primers with an alpha-functionalized acetyl-CoA as the extender unit to generate alpha-functionalized β-keto acyl-CoA. Dehydrogenase converts alpha-functionalized β-keto acyl-CoA to alpha-functionalized β-hydroxy acyl-CoA. Dehydratase converts alpha-functionalized β-hydroxy acyl-CoA to alpha-functionalized enoyl-CoA. Reductase converts alpha-functionalized enoyl-CoA to alpha-functionalized acyl-CoA. The platform can be operated in an iterative manner (i.e. multiple turns) by using the resulting alpha-functionalized acyl-CoA as primer and the aforementioned alpha-functionalized extender unit in subsequent turns of the cycle. Termination pathways acting on any of the four alpha-functionalized CoA thioester intermediates terminate the platform and generate various alpha-functionalized carboxylic acids, alcohols and amines with different β-reduction degree.

Microorganisms are genetically engineered to produce various chemicals for industrial use. The microorganisms are carboxydotrophic acetogens. The microorganisms produce acetyl-CoA using the Wood-Ljungdahl Pathway for fixing CO/CO.sub.2. A reverse beta-oxidation pathway cycle from a microorganism that contains such a group of enzymes is introduced. Additionally, primers and extenders, and/or genes encoding for enzymes that generate primers and extenders may also be introduced. Product synthesis can be effected by improved promoters or enzyme designs that are catalytically more efficient. Similarly, product synthesis may also be improved by deleting competing reactions.