US 10,889,842 B2
Microorganisms for the enhanced production of amino acids and related methods
Renee M. Saville, Mountain View, CA (US); Joshua A. Silverman, Los Altos Hills, CA (US); Eric G. Luning, Sunnyvale, CA (US); Brandon D. Doss, Mountain View, CA (US); Lorraine Joan Giver, Sunnyvale, CA (US); Sol M. Resnick, Encinitas, CA (US); and Drew D. Regitsky, San Francisco, CA (US)
Assigned to CALYSTA, INC., Menlo Park, CA (US)
Filed by CALYSTA, INC., Menlo Park, CA (US)
Filed on Jan. 16, 2015, as Appl. No. 14/599,383.
Claims priority of provisional application 61/928,401, filed on Jan. 16, 2014.
Prior Publication US 2015/0197779 A1, Jul. 16, 2015
Int. Cl. C12N 1/04 (2006.01); C12N 1/32 (2006.01); C12P 13/08 (2006.01); C12P 13/12 (2006.01); C12N 15/52 (2006.01); C12N 1/30 (2006.01)
CPC C12P 13/08 (2013.01) [C12N 1/30 (2013.01); C12N 15/52 (2013.01); C12P 13/12 (2013.01)] 21 Claims
 
1. A recombinant C1 metabolizing microorganism, comprising:
(a) a first heterologous nucleic acid selected from
(i) a heterologous nucleic acid that encodes an L-amino acid biosynthesis enzyme for producing a desired amino acid selected from L-lysine, L-tryptophan, L-methionine, L-cysteine, L-threonine, L-histidine, and L-valine, and
(ii) a heterologous nucleic acid comprising an expression control sequence that is operably linked to a nucleic acid that encodes a native L-amino acid biosynthesis enzyme for producing the desired amino acid, and
(b) a second heterologous nucleic acid that encodes a cache polypeptide that comprises at least 70 amino acid residues in length and comprises the desired amino acid at a level that is 10% or more of the total number of amino acid residues in the cache polypeptide;
wherein the recombinant C1 metabolizing microorganism is capable of converting a natural gas-derived carbon feedstock into the desired L-amino acid at an increased level as compared to the non-genetically engineered C1 metabolizing microorganism,
wherein a δ13C value of the recombinant C1 metabolizing microorganism is less than −30‰, and
wherein
when the desired amino acid is L-lysine, the L-amino acid biosynthesis enzyme of (a) (i) or the native L-amino acid biosynthesis enzyme of (a) (ii) is a lysine biosynthesis enzyme, and the cache polypeptide of (b) comprises lysine at a level that is 10% or more of the total number of amino acid residues in the cache polypeptide of (b),
when the desired amino acid is L-tryptophan, the L-amino acid biosynthesis enzyme of (a) (i) or the native L-amino acid biosynthesis enzyme of (a) (ii) is a tryptophan biosynthesis enzyme, and the cache polypeptide of (b) comprises tryptophan at a level that is 10% or more of the total number of amino acid residues in the cache polypeptide of (b),
when the desired amino acid is L-methionine, the L-amino acid biosynthesis enzyme of (i) or the native L-amino acid biosynthesis enzyme of (ii) is a methionine biosynthesis enzyme, and the cache polypeptide of (b) comprises methionine at a level that is 10% or more of the total number of amino acid residues in the cache polypeptide of (b),
when the desired amino acid is L-cysteine, the L-amino acid biosynthesis enzyme of (i) or the native L-amino acid biosynthesis enzyme of (ii) is a cysteine biosynthesis enzyme, and the cache polypeptide of (b) comprises cysteine at a level that is 10% or more of the total number of amino acid residues in the cache polypeptide of (b),
when the desired amino acid is L-threonine, the L-amino acid biosynthesis enzyme of (i) or the native L-amino acid biosynthesis enzyme of (ii) is a threonine biosynthesis enzyme, and the cache polypeptide of (b) comprises threonine at a level that is 10% or more of the total number of amino acid residues in the cache polypeptide of (b),
when the desired amino acid is L-histidine, the L-amino acid biosynthesis enzyme of (i) or the native L-amino acid biosynthesis enzyme of (ii) is a histidine biosynthesis enzyme, and the cache polypeptide of (b) comprises histidine at a level that is 10% or more of the total number of amino acid residues in the cache polypeptide of (b), and
when the desired amino acid is L-valine, the L-amino acid biosynthesis enzyme of (i) or the native L-amino acid biosynthesis enzyme of (ii) is a valine biosynthesis enzyme, and the cache polypeptide of (b) comprises valine at a level that is 10% or more of the total number of amino acid residues in the cache polypeptide of (b).