US 11,753,663 B2
Microorganisms and methods for enhancing the availability of reducing equivalents in the presence of methanol, and for producing adipate, 6-aminocaproate, hexamethylenediamine or caprolactam related thereto
Anthony P. Burgard, Bellefonte, PA (US); Robin E. Osterhout, San Diego, CA (US); Stephen J. Van Dien, Encinitas, CA (US); Cara Ann Tracewell, Solana Beach, CA (US); Priti Pharkya, San Diego, CA (US); and Stefan Andrae, San Diego, CA (US)
Assigned to Genomatica, Inc., San Diego, CA (US)
Appl. No. 14/652,727
Filed by Genomatica, Inc., San Diego, CA (US)
PCT Filed Dec. 16, 2013, PCT No. PCT/US2013/075287
§ 371(c)(1), (2) Date Jun. 16, 2015,
PCT Pub. No. WO2014/099725, PCT Pub. Date Jun. 26, 2014.
Claims priority of provisional application 61/738,306, filed on Dec. 17, 2012.
Claims priority of provisional application 61/766,620, filed on Feb. 19, 2013.
Prior Publication US 2015/0329885 A1, Nov. 19, 2015
This patent is subject to a terminal disclaimer.
Int. Cl. C12P 17/10 (2006.01); C12P 13/02 (2006.01); C12P 7/24 (2006.01); C12P 19/02 (2006.01); C12P 7/44 (2006.01); C12P 13/00 (2006.01); C12N 9/04 (2006.01); C08G 63/78 (2006.01); C08G 73/02 (2006.01); C08G 69/16 (2006.01); C08G 69/08 (2006.01); C12N 15/52 (2006.01); A23L 29/00 (2016.01)
CPC C12P 17/10 (2013.01) [A23L 29/065 (2016.08); C08G 63/78 (2013.01); C08G 69/08 (2013.01); C08G 69/16 (2013.01); C08G 73/0213 (2013.01); C12N 9/0006 (2013.01); C12N 15/52 (2013.01); C12P 7/24 (2013.01); C12P 7/44 (2013.01); C12P 13/001 (2013.01); C12P 13/005 (2013.01); C12Y 101/01244 (2013.01)] 20 Claims
 
1. A non-naturally occurring microbial organism (NNOMO) comprising:
(A) a methanol metabolic pathway (MMP) consisting of a combination of MMP enzymes (MMPEs) selected from the group consisting of:
(i) a methanol dehydrogenase (EM9), a methylenetetrahydrofolate dehydrogenase (EM3), a methenyltetrahydrofolate cyclohydrolase (EM4) and a formyltetrahydrofolate deformylase (EM5);
(ii) an EM9, an EM3, an EM4, and a formyltetrahydrofolate synthetase (EM6);
(iii) an EM9 and a formaldehyde dehydrogenase (EM11);
(iv) an EM9, a S-(hydroxymethyl)glutathione synthase (EM12), a glutathione-dependent formaldehyde dehydrogenase (EM13), and a S-formylglutathione hydrolase (EM14);
(v) an EM9, an EM13, and an EM14;
(vi) an EM9, a formaldehyde activating enzyme (EM10), an EM3, an EM4, and an EM5; and
(vii) an EM9, an EM10, an EM3, an EM4, and an EM6;
wherein said MMPEs of said MMP are encoded by exogenous nucleic acids;
wherein said MMP enhances the availability of reducing equivalents in the presence of methanol; and
wherein a chemical conversion of said MMP consisting of said combination of MMPEs of group (i) or (ii) comprises spontaneous conversion of formaldehyde to methylene-THF and a chemical conversion of said MMP consisting of the combination of MMPEs of group (vii) comprises spontaneous conversion of formaldehyde to S-hydroxymethylglutathione; and
(B) (i) an adipate pathway (AdiP),
(ii) a 6-aminocaproate (6-ACA) pathway (6-ACAP),
(iii) a hexamethylenediamine (HMDA) pathway (HMDAP), or
(iv) a caprolactam pathway (CapP).