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.

The purpose of the present invention is to provide a method for enhancing the production quantity of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (P(3HB-co-3HH)) having a high fraction of 3-hydroxyhexanoate (3HH) using a saccharide or glycerol as a starting material. The present invention provides: a method for producing a P(3HB-co-3HH) copolymer including performing transformation by homologous recombination of a crotonyl-CoA reductase gene in a chromosome of a recombinant strain of Cupriavidus necator endowed with the ability to produce P(3HB-co-3HH), or performing transformation by introducing an autonomous replication vector in which the crotonyl-CoA reductase gene is incorporated in the aforementioned strain, and cultivating the transformant in a medium containing a saccharide or glycerol as a carbon source; and a method for enhancing the production quantity of the copolymer and/or the fraction of 3HHx in the copolymer.

A recombinant bacterium with a high PHA yield and its construction method provide an engineered microorganism that can be used for the production of polyhydroxyalkanoate (PHA). A recombinant Ralstonia eutropha with a high PHA yield includes a promoter for upregulating a phaJ gene and further includes a mutant phaC gene. The recombinant Ralstonia eutropha can be used for producing 3-hydroxybutyrate-co-3-hydroxyhexanoate (PHBHHx) in 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 microorganisms and methods for the production of polyhydroxybutyrate (PHB) from gaseous substrates. In particular, the invention provides a non-naturally occurring Wood-Ljungdahl microorganism comprising (a) an enzyme that converts acetyl-CoA to acetoacetyl-CoA, (b) an enzyme that converts acetoacetyl-CoA to 3-hydroxybutyryl-CoA, and (c) an enzyme that converts 3-hydroxybutyryl-CoA to polyhydroxybutyrate, and methods related thereto.

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 -keto acyl-CoA. Dehydrogenase converts alpha-functionalized -keto acyl-CoA to alpha-functionalized -hydroxy acyl-CoA. Dehydratase converts alpha-functionalized -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 -reduction degree.

Provided are a method for enhancing the production quantity of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (P(3HB-co-3HH)) having a high fraction of 3-hydroxyhexanoate (3HH) using a saccharide or glycerol as a starting material; a method for producing a P(3HB-co-3HH) copolymer including performing transformation by homologous recombination of a crotonyl-CoA reductase gene in a chromosome of a recombinant strain of Cupriavidus necator endowed with the ability to produce P(3HB-co-3HH), or performing transformation by introducing an autonomous replication vector in which the crotonyl-CoA reductase gene is incorporated in the aforementioned strain, and cultivating the transformant in a medium containing a saccharide or glycerol as a carbon source; and a method for enhancing the production quantity of the copolymer and/or the fraction of 3HHx in the copolymer.

The document provides methods for biosynthesizing isobutene using one or more isolated enzymes such as one or more of a hydratase such as an enzyme classified under EC 4.2.1.- and a decarboxylating thioesterase, or using recombinant host cells expressing one or more such enzymes.

A method of producing polyhydroxyalkanoate (PHA) copolymers comprising both short-chain-length (scl) and medium-chain-length (mcl) subunits, wherein some of the mcl subunits bear reactive vinyl groups, is provided. The method comprises providing cells comprising (i) a PHA synthase (phaC) gene encoding a particular class I poly(3-hydroxyalkanoate) polymerase, and (ii) a phaJ gene encoding a particular (R)-specific enoyl-CoA hydratase, and cultivating said cells in a growth medium comprising an alkenoic acid. In one example, cells grown in the presence of a mixture of decanoic acid and undecen-10-enoic acid yielded a copolymer comprising units of 3-hydroxybutyrate (94.73 mol %), 3-hydroxyhexanoate (1.51 mol %), 3-hydroxyoctanoate (2.33 mol %), 3-hydroxydecanoate (0.69 mol %) and 3-hydroxyhept-6-enoate (0.73 mol %). The presence of acrylic acid as a 13-oxidation inhibitor in the growth medium led to an increase in both the mcl-PHA subunit content and vinyl subunit content. For example, a copolymer containing about 1.2 mol % 3-hydroxyhept-6-enoate and about 0.5 mol % 3-hydroxynon-8-enoate was obtained at a concentration of 40 mM acrylic acid. PHA accumulation of up to 57.2% (w/w of cell dry weight) are reported. The scl-mcl PHAs with subunits bearing reactive vinyl groups are expected to be useful in a variety of applications, for instance for covalently linking bioactive molecules.

The present invention relates to the technical field of microorganisms, and specifically to engineered microorganisms expressing acetoacetyl-CoA reductase variants and methods for increasing the proportion of 3-hydroxyhexanoic acid in PHA. The acetoacetyl-CoA reductase variants and their coding genes provided by the present invention can significantly increase the proportion of 3-hydroxyhexanoic acid in PHA produced by strains; the proportion of 3-hydroxyhexanoic acid in PHA produced by the engineered Ralstonia eutropha constructed utilizing the acetoacetyl-CoA reductase variants and their coding genes provided by the present invention is significantly increased, which provides new genes and strain resources for strains producing poly(3-hydroxybutyrate-co-3-10 the development of engineered hydroxyhexanoate) with high proportion of 3-hydroxyhexanoic acid.