Production of fatty acids & derivatives thereof

Abstract

Compositions and methods for production of fatty alcohols using recombinant microorganisms are provided as well as fatty alcohol compositions produced by such methods.

1 . A method of producing a fatty alcohol composition in a recombinant microorganism, comprising the steps of:
 (a) genetically engineering a microorganism to comprise a nucleic acid sequence encoding a polypeptide having acetyl-CoA carboxylase activity (EC 6.4.1.2), and a nucleic acid sequence encoding a polypeptide having fatty alcohol forming activity, resulting in a recombinant microorganism;   (b) culturing the recombinant microorganism in a culture medium containing a carbon source under conditions effective to overexpress the acetyl-CoA carboxylase polypeptide and the polypeptide having fatty alcohol forming activity, wherein a fatty alcohol composition is produced by said cultured recombinant microorganism; and   (c) optionally recovering the fatty alcohol composition from the cell culture.   
     
     
         2 . The method of  claim 1 , further comprising genetically engineering said microorganism to comprise at least one nucleic acid sequence encoding a polypeptide having thioesterase activity, wherein said thioesterase polypeptide is expressed. 
     
     
         3 . The method of  claim 1 , wherein said polypeptide having fatty alcohol forming activity is (i) a fatty alcohol forming acyl-CoA reductase (FAR, EC 1.1.1.*), or (ii) an acyl-CoA reductase (EC 1.2.1.50) and an alcohol dehydrogenase (EC 1.1.1.1). 
     
     
         4 . The method of  claim 2 , wherein said polypeptide having fatty alcohol forming activity is (i) a fatty alcohol forming acyl-CoA reductase (FAR, EC 1.1.1.*), or (ii) an acyl-CoA reductase (EC 1.2.1.50) and an alcohol dehydrogenase (EC 1.1.1.1). 
     
     
         5 . The method of  claim 4 , wherein said polypeptide having fatty alcohol forming activity is a fatty alcohol forming acyl-CoA reductase. 
     
     
         6 . The method of  claim 1 , wherein said the fatty alcohol composition comprises one or more of saturated or unsaturated C12, C14 or C16 fatty alcohols. 
     
     
         7 . A recombinant microorganism comprising:
 (a) a nucleic acid sequence encoding a branched chain alpha-keto acid dehydrogenase 60 (Bkd) operon including branched-chain α-keto acid decarboxylase α and β subunits (E1α/β), a dihydrolipoyl transacylase component (E2), and a dihydrolipoyl dehydrogenase component (E3); and   (b) a nucleic acid sequence encoding a β-ketoacyl-ACP synthase III protein   (FabH, EC 2.3.1.41) with specificity for a branched chain acyl CoA molecule, wherein at least one nucleic acid sequence according to (a) or (b) is exogenous to the recombinant microorganism and wherein the recombinant microorganism produces a branched fatty acid derivative when cultured in the presence of a carbon source under conditions effective to express the nucleic acid sequences according to (a) and (b).   
     
     
         8 . The recombinant microorganism according to  claim 7 , wherein the nucleic acid sequence encoding the FabH protein with specificity for a branched chain acyl CoA molecule is exogenous to the recombinant microorganism and the expression of a FabH endogenous to the recombinant microorganism and lacking specificity for a branched chain acyl CoA molecule is attenuated. 
     
     
         9 . The recombinant microorganism according to  claim 7 , further comprising a nucleic acid sequence encoding at least one polypeptide having thioesterase activity. 
     
     
         10 . The recombinant microorganism according to  claim 9 , further comprising a nucleic acid sequence encoding a polypeptide having fatty alcohol forming activity. 
     
     
         11 . The recombinant microorganism according to  claim 10 , wherein said polypeptide having fatty alcohol forming activity is (i) a fatty alcohol forming acyl-CoA reductase (FAR, EC 1.1.1.*), or (II) acyl-CoA reductases (EC 1.2.1.50) and alcohol dehydrogenase (EC 1.1.1.1). 
     
     
         12 . The recombinant microorganism according to  claim 11 , wherein said polypeptide having fatty alcohol forming activity is a fatty alcohol forming acyl-CoA reductase. 
     
     
         13 . A recombinant microorganism culture, comprising:
 the recombinant microorganism according to  claim 11  and a fermentation medium comprising a carbon source.   
     
     
         14 . A branched fatty alcohol composition produced by the recombinant microorganism culture according to  claim 13 , wherein said fatty alcohol composition comprises one or more of saturated or unsaturated C12, C14 and C16 fatty alcohols. 
     
     
         15 . A branched fatty alcohol composition obtained from the supernatant of the recombinant microorganism culture of  claim 13  wherein the fatty alcohol composition comprises C 12  and C 14  fatty alcohols. 
     
     
         16 . A branched fatty alcohol composition obtained from the supernatant of the recombinant microorganism culture of  claim 13 , wherein the fatty alcohol composition comprises unsaturated fatty alcohols. 
     
     
         17 . A branched fatty alcohol composition obtained from the supernatant of the recombinant microorganism culture of  claim 13 , wherein the fatty alcohol composition comprises saturated fatty alcohols. 
     
     
         18 . A method of producing a branched fatty alcohol composition in a recombinant microorganism, comprising the steps of:
 (a) obtaining a genetically engineered recombinant microorganism according to  claim 11 ;   (b) culturing the recombinant microorganism in a culture medium containing a carbon source under conditions effective to express said: (i) Bkd operon; (ii) FabH;   (iii) a nucleic acid sequence encoding a polypeptide having fatty alcohol forming activity; and (iv) nucleic acid sequence encoding a polypeptide having thioesterase activity; and   (c) optionally recovering the branched fatty alcohol composition from the cell culture.   
     
     
         19 . The method of  claim 18 , wherein said polypeptide having fatty alcohol forming activity is (i) a fatty alcohol forming acyl-CoA reductase (FAR, EC 1.1.1.*), or (II) an acyl-CoA reductase (EC 1.2.1.50) and an alcohol dehydrogenase (EC 1.1.1.1). 
     
     
         20 . The recombinant microorganism according to  claim 19 , wherein said polypeptide having fatty alcohol forming activity is a fatty alcohol forming acyl-CoA reductase.