Methods and organisms for utilizing synthesis gas or other gaseous carbon sources and methanol

Abstract

The invention provides a non-naturally occurring microbial organism having an acetyl-CoA pathway and the capability of utilizing syngas or syngas and methanol. In one embodiment, the invention provides a non-naturally occurring microorganism, comprising one or more exogenous proteins conferring to the microorganism a pathway to convert CO, CO2 and/or H2 to acetyl-coenzyme A (acetyl-CoA), methyl tetrahydrofolate (methyl-THF) or other desired products, wherein the microorganism lacks the ability to convert CO or CO2 and H2 to acetyl-CoA or methyl-THF in the absence of the one or more exogenous proteins. For example, the microbial organism can contain at least one exogenous nucleic acid encoding an enzyme or protein in an acetyl-CoA pathway. The microbial organism is capable of utilizing synthesis gases comprising CO, CO2 and/or H2, alone or in combination with methanol, to produce acetyl-CoA. The invention additionally provides a method for producing acetyl-CoA, for example, by culturing an acetyl-CoA producing microbial organism, where the microbial organism expresses at least one exogenous nucleic acid encoding an acetyl-CoA pathway enzyme or protein in a sufficient amount to produce acetyl-CoA, under conditions and for a sufficient period of time to produce acetyl-CoA.

1 - 191 . (canceled) 
     
     
         192 . A non-naturally occurring microorganism, comprising an exogenous nucleic acid encoding a carbon monoxide dehydrogenase, wherein said carbon monoxide dehydrogenase generates reducing equivalents that are electrons, hydrogen atoms, or both, from carbon monoxide (CO), or from a mixture of CO and hydrogen (H 2 ), wherein the microorganism lacks the ability to convert CO and H 2  to acetyl-CoA in the absence of carbon monoxide dehydrogenase, and wherein the non-naturally occurring microorganism produces an amount of acetyl-CoA that is greater than an amount of acetyl-CoA produced in the same microorganism without the carbon monoxide dehydrogenase. 
     
     
         193 . The microorganism of  claim 192  wherein said microbial organism further comprises an exogenous nucleic acid encoding a ferredoxin. 
     
     
         194 . The microorganism of  claim 192  wherein said microbial organism further comprises an exogenous nucleic acid encoding a pyruvate ferredoxin oxidoreductase. 
     
     
         195 . The microorganism of  claim 192  wherein said microbial organism further comprises an exogenous nucleic acid encoding a hydrogenase, wherein said hydrogenase generates reducing equivalents from H 2 . 
     
     
         196 . The microorganism of  claim 192  wherein said microbial organism further comprises an exogenous nucleic acid encoding a formate dehydrogenase. 
     
     
         197 . The microorganism of  claim 192  wherein said microbial organism further comprises an exogenous nucleic acid encoding a hydrogenase, wherein said hydrogenase generates reducing equivalents from H 2 . 
     
     
         198 . The microorganism of  claim 197  wherein said hydrogenase is 3-hydroxyacyl dehydrogenase 
     
     
         199 . The non-naturally occurring bacterial microorganism of  claim 198  wherein the 3-hydroxyacyl dehydrogenase is 3-hydroxybutyryl-CoA dehydrogenase. 
     
     
         200 . The non-naturally occurring bacterial microorganism of  claim 199  wherein the 3-hydroxybutyryl-CoA dehydrogenase is encoded by a gene selected from the group consisting of  C. acetobutylicum  hbd,  C. beijerinckii  hbd,  Metallosphaera sedula  Msed_1423 , Metallosphaera sedula  Msed_0399 , Metallosphaera sedula  Msed_0389, and  Metallosphaera sedula  Msed_1993. 
     
     
         201 . The non-naturally occurring bacterial microorganism of  claim 192  further comprising an enzyme that coverts 3-hydroxybutyryl-CoA to 4-hydroxybutyryl-CoA or crotonyl-CoA. 
     
     
         202 . The non-naturally occurring bacterial microorganism of  claim 201  wherein the enzyme that coverts 3-hydroxybutyryl-CoA to 4-hydroxybutyryl-CoA or crotonyl-CoA is enoyl-CoA hydratase. 
     
     
         203 . The non-naturally occurring bacterial microorganism of  claim 192  wherein said carbon monoxide dehydrogenase is encoded by a gene selected from the group consisting of  Moorella thermoacetica  acsA and  Methanosarcina acetivorans  acsA. 
     
     
         204 . The non-naturally occurring bacterial microorganism of  claim 192  selected from  Escherichia coli, Klebsiella oxytoca, Anaerobiospirillum succiniciproducens, Actinobacillus succinogenes, Mannheimia succiniciproducens, Rhizobium etli, Bacillus subtilis, Corynebacterium glutamicum, Gluconobacter oxydans, Zymomonas mobilis, Lactococcus lactis, Lactobacillus plantarum, Streptomyces coelicolor, Clostridium acetobutylicum, Pseudomonas fluorescens , and  Pseudomonas putida.