Monsanto Company

Steven Charles Slater

Ganesh M Kishore

Timothy A Mitsky

David M Stark

Stephen Rogers Padgette

Kenneth James Gruys

Genes and methods for optimizing levels of substrates employed in the biosynthesis of copolymers of 3-hydroxybutyrate (3HB) and 3-hydroxyvalerate (3HV) in plants and bacteria via manipulation of normal metabolic pathways using recombinant DNA techniques are provided. This is achieved through the use of a variety of wild-type and/or deregulated enzymes involved in the biosynthesis of aspartate family amino acids, and wild-type or deregulated forms of enzymes, such as threonine deaminase, involved in the conversion of threonine to P(3HB-co-3HV) copolymer endproduct. By these methods, enhanced levels of threonine, .alpha.-ketobutyrate, propionate, propionyl-CoA, .beta.-ketovaleryl-CoA, and .beta.-hydroxyvaleryl-CoA are produced. Also provided are methods for the biological production of P(3HB-co-3HV) copolymers in plants and bacteria utilizing propionyl-CoA produced through a variety of engineered metabolic pathways. Introduction into plants and bacteria of an appropriate .beta.-ketothiolase, .beta.-ketoacyl-CoA reductase, and PHA synthase, alone or in combination with various enzymes involved in asparate family amino acid biosynthesis and the conversion of threonine to PHA copolymer precursors, will permit these organisms to produce P(3HB-co-3HV) copolymers.

Methods of optimizing substrate pools and biosynthesis of

Genes and methods for optimizing levels of substrates employed in the biosynthesis of copolymers of 3-hydroxybutyrate (3HB) and 3-hydroxyvalerate (3HV) in plants and bacteria via manipulation of normal metabolic pathways using recombinant DNA techniques are provided. This is achieved through the use of a variety of wild-type and/or deregulated enzymes involved in the biosynthesis of aspartate family amino acids, and wild-type or deregulated forms of enzymes, such as threonine deaminase, involved in the conversion of threonine to P(3HB-co-3HV) copolymer end product. By these methods, enhanced levels of threonine, .alpha.-ketobutyrate, propionate, propionyl-CoA, .beta.-ketovaleryl-CoA, and .beta.-hydroxyvaleryl-CoA are produced. Also provided are methods for the biological production of P(3HB-co-3HV) copolymers in plants and bacteria utilizing propionyl-CoA produced through a variety of engineered metabolic pathways. Introduction into plants and bacteria of an appropriate .beta.-ketothiolase, .beta.-ketoacyl-CoA reductase, and PHA synthase, alone or in combination with various enzymes involved in asparate family amino acid biosynthesis and the conversion of threonine to PHA copolymer precursors, will permit these organisms to produce P(3HB-co-3HV) copolymers.

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Chesterfield, MO, United States of America

Steven Charles Slater