BIOLOGICAL METHODS FOR PREPARING ADIPIC ACID
The technology relates in part to biological methods for producing adipic acid and engineered microorganisms capable of such production.
1 - 213 . (canceled)
214 . An engineered yeast comprising modifications that add or increase a hexanoate synthase activity, a monooxygenase activity, and a monooxygenase reductase activity, which yeast is capable of producing a diacid from a sugar.
215 . The engineered yeast of claim 214 , which comprises increased copies of a polynucleotide encoding a polypeptide having the hexanoate synthase activity, the monooxygenase activity, or the monooxygenase reductase activity.
216 . The engineered yeast of claim 214 , which comprises a heterologous promoter and/or 5′ UTR in genomic DNA in functional connection with a polynucleotide encoding a polypeptide having the hexanoate synthase activity, the monooxygenase activity, or the monooxygenase reductase activity.
217 . The engineered yeast of claim 214 , wherein the hexanoate synthase activity is provided by a heterologous polypeptide.
218 . The engineered yeast of claim 217 , wherein the heterologous polypeptide is encoded by a hexonate synthase subunit alpha (HEXA) gene, hexanoate synthase subunit beta (HEXB) gene, or a hexonate synthase subunit alpha (HEXA) gene and hexanoate synthase subunit beta (HEXB) gene.
219 . The engineered yeast of claim 218 , wherein the HEXA gene, HEXB gene, or HEXA gene and HEXB gene are from an Aspergillus spp. fungus.
220 . The engineered yeast of claim 219 , wherein the Aspergillus spp. fungus is Aspergillus parasiticus.
221 . The engineered yeast of claim 214 , which includes an increased number of copies of an endogenous polynucleotide that encodes a polypeptide having the monooxygenase activity.
222 . The engineered yeast of claim 221 , wherein the endogenous polynucleotide encodes a CYP52A15 polypeptide, CYP52A16 polypeptide or CYP52A19 polypeptide.
223 . The engineered yeast of claim 214 , which comprises a heterologous promoter in genomic DNA in functional connection with an endogenous polynucleotide that encodes a polypeptide having the monooxygenase activity.
224 . The engineered yeast of claim 223 , wherein the endogenous polynucleotide encodes a CYP52A15 polypeptide, CYP52A16 polypeptide or CYP52A19 polypeptide.
225 . The engineered yeast of claim 214 , which includes an increased number of copies of an endogenous polynucleotide that encodes a polypeptide having the monooxygenase reductase activity.
226 . The engineered yeast of claim 225 , wherein the endogenous polynucleotide encodes a CPRA polypeptide or CPRB polypeptide.
227 . The engineered yeast of claim 214 , which comprises a heterologous promoter in genomic DNA in functional connection with an endogenous polynucleotide that encodes a polypeptide having the monooxygenase reductase activity.
228 . The engineered yeast of claim 227 , wherein the endogenous polynucleotide encodes a CPRA polypeptide or CPRB polypeptide.
229 . The engineered yeast of claim 214 , which is a Candida spp. yeast.
230 . The engineered yeast of claim 214 , which is a Yarrowia spp. yeast.
231 . The engineered yeast of claim 230 , wherein the Yarrowia spp. yeast is Yarrowia lipolytica.
232 . The engineered yeast of claim 214 , wherein the diacid is adipic acid.