Biological methods for preparing a fatty dicarboxylic acid

Abstract

Provided are engineered microorganisms capable of producing fatty dicarboxylic acids and products expressed by such microorganisms. Also provided are biological methods for producing fatty dicarboxylic acids.

1 .- 98 . (canceled) 
     
     
         99 . A method for producing a desired diacid by a yeast from a feedstock toxic to the yeast, comprising: (a) contacting a genetically modified yeast in culture with a feedstock not substantially toxic to the yeast, thereby performing an induction; and (b) contacting the yeast after the induction in (a) with a feedstock toxic to the yeast, whereby the desired diacid is produced by the yeast from the feedstock toxic to the yeast in an amount greater than the amount of the diacid produced from the feedstock toxic to the yeast when the induction is not performed. 
     
     
         100 . The method of  claim 99 , wherein the feedstock not substantially toxic to the yeast has the same number of carbons as the feedstock toxic to the yeast. 
     
     
         101 . The method of  claim 99 , wherein the feedstock not substantially toxic to the yeast has a different number of carbons compared to the feedstock toxic to the yeast. 
     
     
         102 . The method of  claim 99 , wherein the feedstock not substantially toxic to the yeast comprises a fatty acid methyl ester. 
     
     
         103 . The method of  claim 99 , wherein the feedstock not substantially toxic to the yeast comprises a free fatty acid. 
     
     
         104 . The method of  claim 99 , wherein the feedstock not substantially toxic to the yeast comprises more than twelve carbons. 
     
     
         105 . A diacid produced by the method of  claim 99 . 
     
     
         106 .- 176 . (canceled) 
     
     
         177 . The method of  claim 102 , wherein the fatty acid ethyl ester is methyl myristate. 
     
     
         178 . The method of  claim 99 , wherein the feedstock toxic to the yeast is a fatty acid methyl ester. 
     
     
         179 . The method of  claim 103 , wherein the free fatty acid is decane. 
     
     
         180 . The method of  claim 178 , wherein the fatty acid methyl ester is methyl decanoate. 
     
     
         181 . The method of  claim 99 , wherein the feedstock toxic to the yeast is a free fatty acid. 
     
     
         182 . The method of  claim 181 , wherein the free fatty acid is lauric acid. 
     
     
         183 . The method of  claim 99 , wherein the genetically modified yeast comprises at least one genetic modification that substantially blocks beta oxidation activity. 
     
     
         184 . The method of  claim 99 , wherein the genetically modified yeast comprises at least one genetic modification at least one genetic modification that increase at least one activity selected from the group consisting of: monooxygenase activity, monooxygenase reductase activity, thioesterase activity, acyltransferase activity, isocitrate dehydrogenase activity, glyceraldehyde-3-phosphate dehydrogenase activity, glucose-6-phosphate dehydrogenase activity, acyl-coA oxidase activity, fatty alcohol oxidase activity, acyl-CoA hydrolase activity, alcohol dehydrogenase activity, peroxisomal biogenesis factor activity, fatty aldehyde dehydrogenase activity, CTF, UTR, FAT1. 
     
     
         185 . The method of  claim 99 , wherein the genetically modified yeast comprises at least one genetic modification that decreases MIG1 activity, as compared to corresponding yeast not comprising said genetic modification. 
     
     
         186 . The method of  claim 184 , wherein the genetically modified yeast comprises at least one genetic modification that increases monooxygase activity selected from the group consisting of: a CYP52A12 monooxygenase activity, CYP52A13 monooxygenase activity, CYP52A14 monooxygenase activity, CYP52A15 monooxygenase activity, CYP52A16 monooxygenase activity, CYP52A17 monooxygenase activity, CYP52A18 monooxygenase activity, CYP52A19 monooxygenase activity, CYP52A20 monooxygenase activity, CYP52D2 monooxygenase activity and BM3 monooxygenase activity. 
     
     
         187 . The method of  claim 184 , wherein the genetically modified yeast comprises at least one genetic modification that increases glucose-6-phosphate dehydrogenase activity selected from the group consisting of: a ZWF1 glucose-6-phosphate dehydrogenase activity and ZWF2 glucose-6-phosphate dehydrogenase activity. 
     
     
         188 . The method of  claim 184 , wherein the genetically modified yeast comprises at least one genetic modification that increases fatty alcohol oxidase activity selected from the group consisting of: FAO1 fatty alcohol oxidase activity, FA02A fatty alcohol oxidase activity, FA02B fatty alcohol oxidase activity, FAO13 fatty alcohol oxidase activity, FAO17 fatty alcohol oxidase activity, FA018 fatty alcohol oxidase activity and FA020 fatty alcohol oxidase activity. 
     
     
         189 . The method of  claim 184 , wherein the genetically modified yeast comprises at least one genetic modification that increases alcohol dehydrogenase activity selected from the group consisting of: ADH1 alcohol dehydrogenase activity, ADH2 alcohol dehydrogenase activity, ADH3 alcohol dehydrogenase activity, ADH4 alcohol dehydrogenase activity, ADH5 alcohol dehydrogenase activity, ADH7 alcohol dehydrogenase activity, ADH8 alcohol dehydrogenase activity and SFA alcohol dehydrogenase activity.