Production of fatty acid derivatives

Abstract

Methods and compositions for producing fatty acid derivatives, for example, fatty esters, and commercial fuel compositions comprising fatty acid derivatives are described.

1 . A method of producing a fatty acid derivative, the method comprising culturing a host cell in the presence of a carbon source, wherein the host cell is genetically engineered to express or overexpress a gene encoding an ester synthase, and isolating the fatty acid derivative. 
     
     
         2 . The method of  claim 1 , further comprising culturing the host cell in the presence of an alcohol. 
     
     
         3 . The method of  claim 1 , wherein the ester synthase gene is obtained from  Acidobacteria bacterium, Acidothermus cellulolyticus, Acinetobacter baumannii, Acinetobacter baylyi, Acinetobacter  sp.,  Acinetobacter baumannii, Acinetobacter  sp.,  Aeromonas hydrophila, Aeromonas salmonicida, Alcaligenes europhus, Alcanivorax borkumensis, Alcanivorax jadensis, Alteromonas macleodii, Anaeromyxobacter dehalogenans, Anaeromyxobacter, Anaeromyxobacter  sp.,  Arabidopsis thaliana, Bradyrhizobium japonicum, Cryptococcus curvatus, Erythrobacter litoralis, Erythrobacter  sp.,  Frankia  sp.,  Fundibacter jadensis, gamma proteobacterium, Hahella chejuensis, Homo sapiens, Janibacter  sp.,  Limnobacter  sp., marine gamma proteobacterium,  Marinobacter algicola, Marinobacter aquaeolei, Marinobacter hydrocarbinoclasticus, Marinobacter  sp.,  Methylibium petroleiphilum, Microscilla marina, Moritella  sp.,  Mortierella alpina, Mus musculus, Mycobacterium abscessus, Mycobacterium avium, Mycobacterium avium, Mycobacterium bovis, Mycobacterium gilvum, Mycobacterium leprae, Mycobacterium marinum, Mycobacterium smegmatis, Mycobacterium  sp.,  Mycobacterium tuberculosis, Mycobacterium ulcerans, Mycobacterium vanbaalenii, Myxococcus xanthus, Natronomonas pharaonis, Nocardia farcinica, Nocardioides sp.,  Photobacterium profundum, Plesiocystis pacifica, Polaromonas naphthalenivorans, Polaromonas  sp.,  Psudomonas aeruginosa, Psychrobacter arcticus, Psychrobacter cryohalolentis, Psychrobacter  sp.,  Reinekea  sp.,  Rhodococcus opacus, Rhodoferax ferrireducens, Rhodococcus  sp.,  Rhodoferax ferrireducens, Roseiflexus  sp.,  Roseiflexus castenholzii, Saccharomyces cerevisiae, Saccharopolyspora erythraea, Salinibacter ruber, Simmodsia chinensis, Solibacter usitatus, Sphingopyxis alaskensis, Stigmatella aurantiaca, Streptomyces avermitilis, Streptomyces coelicolor, Streptomyces griseus, Tenacibaculum  sp., or  Ustilago maydis.    
     
     
         4 . The method of  claim 3 , wherein the gene encoding an ester synthase is wax/dgat, a gene encoding a wax synthase, a gene encoding a bifunctional ester synthase/acyl-CoA:diacylglycerol acyltransferase, ES9, ES8, DSM 8798, AtfA 1, or AtfA2. 
     
     
         5 . The method of  claim 1 , wherein the gene encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:18, 24, 25, or 26. 
     
     
         6 . The method of  claim 1 , wherein the gene encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:18, 24, 25, or 26, with one or more amino acid substitutions, additions, insertions or deletions. 
     
     
         7 . The method of  claim 6 , wherein the polypeptide comprises the amino acid sequence of SEQ ID NO:18, with a substitution at amino acid residue 395. 
     
     
         8 . The method of  claim 1 , wherein the host cell is genetically engineered to express, relative to a wild type host cell, a decreased level of at least one gene encoding a fatty acid degradation enzyme. 
     
     
         9 . The method of  claim 8 , wherein the fatty acid degradation enzyme gene encodes an acyl-CoA synthase. 
     
     
         10 . The method of  claim 1 , wherein the host cell is genetically engineered to express, relative to a wild type host cell, a decreased level of at least one thioesterase gene. 
     
     
         11 . The method of  claim 1 , wherein the host cell is genetically engineered to express, relative to a wild type host cell, a decreased level of at least one gene encoding an acyl-CoA dehydrogenase. 
     
     
         12 . The method of  claim 1 , wherein the host cell is genetically engineered to express, relative to a wild type host cell, a decreased level of a gene encoding an outer membrane protein receptor. 
     
     
         13 . The method of  claim 1 , wherein the host cell is a mammalian cell, a plant cell, an insect cell, a yeast cell, a fungus cell, a filamentous fungi cell, a cyanobacterial cell, or a bacterial cell. 
     
     
         14 . A fatty acid derivative produced by the method of  claim 1 . 
     
     
         15 . A fatty ester produced by the method of  claim 1 . 
     
     
         16 . A genetically engineered microorganism comprising at least one gene encoding an ester synthase, wherein the gene encoding the ester synthase is overexpressed relative to a wild type ester synthase, and wherein the microorganism produces an increased level of a fatty ester relative to a wild-type microorganism. 
     
     
         17 . A genetically engineered microorganism comprising an exogenous control sequence stably incorporated into the genomic DNA of the microorganism upstream of a gene encoding an ester synthase, wherein the microorganism produces an increased level of a fatty ester relative to a wild-type microorganism. 
     
     
         18 . The genetically engineered microorganism of  claim 16 , wherein the microorganism is genetically engineered to express, relative to a wild type microorganism, a decreased level of at least one gene encoding a fatty acid degradation enzyme. 
     
     
         19 . The genetically engineered microorganism of  claim 16 , wherein the microorganism is genetically engineered to express, relative to a wild type microorganism cell, a decreased level of at least one thioesterase gene. 
     
     
         20 . The genetically engineered microorganism of  claim 16 , wherein the microorganism is genetically engineered to express, relative to a wild type microorganism, a decreased level of at least one gene encoding an acyl-CoA dehydrogenase. 
     
     
         21 . The genetically engineered microorganism of  claim 16 , wherein the microorganism is genetically engineered to express, relative to a wild type microogranism, a decreased level of a gene encoding an outer membrane protein receptor. 
     
     
         22 . The genetically engineered microorganism of  claim 16 , selected from a Gram-negative or a Gram-positive bacterium. 
     
     
         23 . A method of producing a fatty acid derivative comprising culturing the genetically engineered microorganism of  claim 16 , in the presence of an alcohol. 
     
     
         24 . The method of  claim 23 , wherein the alcohol is ethanol, methanol, propanol, or butanol. 
     
     
         25 . A fatty acid derivative produced by the method of  claim 23 . 
     
     
         26 . The fatty acid derivative of  claim 25 , wherein the fatty acid derivative is a fatty ester. 
     
     
         27 . A biofuel composition comprising the fatty acid derivative of  claim 14 . 
     
     
         28 . The biofuel composition of  claim 27 , wherein the composition has a cloud point of about 5° C. or lower, of about 0° C. or lower, or of about −4° C. or lower. 
     
     
         29 . The biofuel composition of  claim 27 , wherein the composition has a simulated distillation T90 of about 360° C. or lower, or of about 358° C. or lower. 
     
     
         30 . The biofuel composition of  claim 27 , wherein the composition has a sulfur content of about 15 ppm or less, of about 12 ppm or less, or of about 10.7 ppm or less. 
     
     
         31 . The biofuel composition of  claim 27 , wherein the composition has a Karl Fischer moisture content of about 0.1 wt. % or less, of about 0.08 wt. % or less, or of about 0.06 wt. % or less. 
     
     
         32 . The biofuel composition of  claim 27 , wherein the composition has a total acid number of about 0.50 mg KOH/g or less, of about 0.20 mg KOH/g or less, or of about 0.15 mg KOH/g or less. 
     
     
         33 . The biofuel composition of  claim 27 , wherein the composition has a combined calcium and magnesium content of about 5 ppm or less, of about 2 ppm or less, or of about 0.5 ppm or less. 
     
     
         34 . The biofuel composition of  claim 27 , wherein the composition has a combined sodium and potassium content of about 5 ppm or less, of about 2.5 ppm or less, or of about 1.6 ppm or less. 
     
     
         35 . The biofuel composition of  claim 27 , wherein the composition has a specific mass at 20° C. of about 850 to about 900 kg/m 3 , of about 860 to about 890 kg/m 3 , or of about 870 to about 880 kg/m 3 . 
     
     
         36 . The biofuel composition of  claim 27 , wherein the composition has a kinematic viscosity at 40° C. of about 3 to about 6 mm 2 /s, of about 3.2 to about 5 mm 2 /s, or of about 3.5 to about 4 mm 2 /s. 
     
     
         37 . The biofuel composition of  claim 27 , wherein the composition has a water content of about 500 mg/kg or less, of about 480 mg/kg or less, or of about 450 mg/kg or less. 
     
     
         38 . The biofuel composition of  claim 27 , wherein the composition has a total contamination level of about 24 mg/kg or less, of about 20 mg/kg or less, of about 10 mg/kg or less, or of about 5 mg/kg or less. 
     
     
         39 . The biofuel composition of  claim 27 , wherein the composition has a flash point of about 100° C. or higher, of about 110° C. or higher, or of about 120° C. or higher. 
     
     
         40 . The biofuel composition of  claim 27 , wherein the composition has a ester content of about 96.5 wt. % or more, or of about 97 wt. % or more. 
     
     
         41 . The biofuel composition of  claim 27 , wherein the composition has a carbon residue number of about 0.05 wt. % or less. 
     
     
         42 . The biofuel composition of  claim 27 , wherein the composition has a sulfated ash level of 0.02 wt. % or less, or of 0.01 wt. % or less. 
     
     
         43 . The biofuel composition of  claim 27 , wherein the composition has a phosphorus level of about 10 mg/kg or less, of about 5 mg/kg or less, or of about 1 mg/kg or less. 
     
     
         44 . The biofuel composition of  claim 27 , wherein the composition has a copper corrosion score of 1. 
     
     
         45 . The biofuel composition of  claim 27 , wherein the composition has a cold filter plugging point of about 19° C. or lower, of about 10° C. or lower, or of about 0° C. or lower. 
     
     
         46 . The biofuel composition of  claim 27 , wherein the composition has a free glycerol level of about 0.02 wt. % or less. 
     
     
         47 . The biofuel composition of  claim 27 , wherein the composition has a total glycerol level of about 0.25 wt. % or less, of about 0.15 wt. % or less, of about 0.10 wt. % or less, or of about 0.05 wt. % or less. 
     
     
         48 . The biofuel composition of  claim 27 , wherein the composition has a monoacylglycerol level of about 0.02 wt. % or less. 
     
     
         49 . The biofuel composition of  claim 27 , wherein the composition has a methanol or ethanol level of about 0.2 wt. % or less, of about 0.1 wt. % or less, or of about 0.02 wt. % or less. 
     
     
         50 . The biofuel composition of  claim 27 , wherein the composition has an iodine number of about 65 g/100 g or less. 
     
     
         51 . The biofuel composition of  claim 27 , wherein the composition has an oxidation stability at 110° C. of about 6 hours or longer, of about 9 hours or longer, or of about 11 hours or longer.